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PANTHER User Manual

1. E Root Folder os Sample 2 50 O 2 sample 1 50 O 2 sample 124 EE uniProtFly 7035 O candidateGene 3 20 2006 59 O 2 dme gene info for Tim 220 Figure 4 3 Choose a list from the workspace 4 1 1 2 Modify reference gene list and proceed Once a list is uploaded it appears on the right side of the page You can delete a list by clicking the x icon in front of the list name Figure 4 4 If multiple IDs are mapped to the same PANTHER ID the tool will only choose one unique ID When all list s are uploaded click the Finished selecting lists button Figure 4 4 The next page allows you to do the following Figure 4 5 CHAPTER 4 PANTHER TOOLS 45 e Modify your selected gene by clicking the Select list s button e Select a reference gene list There are 3 default reference list human mouse rat whole genome list You can click the Select reference list button to upload your own list e Select a type of classification molecular function biological process cellular component protein and pathway you would like to analyze UPLOAD OR SELECT LIST FROM YOUR WORKSPACE Upload list Uploaded and selected lists List type Please select list type B genexpSample_1 ESSA Upload list Browse amonat Waminga There are duplicate IDs in the file The unique set of IDs will be used If there are redundant IDs only the first will be used in the analy
2. Figure 3 5 An example showing selections using Prowler 3 3 Batch ID Search Batch ID search is a unique functionality that you can leverage expert curated PANTHER knowledgebase to classify multiple genes of your interest This is how it works PAN THER library contains genes and proteins from UniProt Refseq ENSEMBL and Model Organism Databases Each gene belongs to a protein subfamily Many subfamilies have been manually curated with ontology terms and therefore the sequence will have the same classifications CHAPTER 3 PANTHER BASICS 37 as the subfamily that it belongs to Using the IDmapping mechanism developed by UniProt each gene is mapped to multiple commonly used IDs including Entrez GeneID and GI Gene Symbol etc referred to as PANTHER Supported IDs When you submit a list of IDs the system will automatically search through all the supported IDs The mapped one will carry over all the PANTHER classifications Below is a brief description of how you can use the tool Figure 3 6 1 Enter your IDs There are two ways to enter your IDs e Type or paste a list of IDs directly to the box e Use the Browse button to upload a list file Batch ID Search Enter IDs separate IDs by a space or comma supported IDs ED Upload IDs Browse file format 1D List Select File Type P Previously exported text search results Result page Genes O Transcripts Proteins Select datasets v Homo sapi
3. item 21 and 22 respectively in Figure 1 1 on the home page allows you to retrieve quick results from our system Section 3 1 Home Browse Genes and orthologs Trees and HMMs Pathways Ontologies Tools Workspace Enter your Email KEYWORD SEARCH SEQUENCE SEARCH Enter a protein sequence Sequence query limits Protein 50kb The PANTHER Protein ANalysis THrough Evolutionary Relationships Classification System is a unique resource that classifies genes by their functions using published scientific experimental evidence and evolutionary relationships to predict function even in the absence of direct experimental evidence Proteins are classified by expert biologists according to gt Gene families and subfamilies including annotated phylogenetic trees Gene Ontology classes molecular function biological process cellular component PANTHER Protein Classes gt Pathways including diagrams PANTHER is part of the Gene Ontology Reference Genome Project PANTHER is supported by a research grant from the National Institute of General Medical Sciences grant GM081084 and maintained by the Thomas lab at the University of Southern California About ContactUs System Requirements Privacy Policy Disclaimer Copyright 2010 Paul Thomas All Rights Reserved Figure 1 1 The PANTHER home page 11 All B La What can do on 2 21 he site E PANTHER Pathways arae
4. 12 keyword search 3 6 32 advanced 33 simple 32 Ladder bottom 29 Ladder original 29 Ladder top 29 LDO see least diverge ortholog leaf node 28 least diverged ortholog 17 LOD see least diverge ortholog login see registration Mann Whitney test 48 50 51 match state 30 31 Model Organism Database 6 molecular function 9 12 16 18 34 MSA 28 30 multiple sequence alignment 28 30 ND 17 NIC see Number of independent counts Number of independent counts 55 ontology 3 34 35 58 INDEX ontology home page 9 ortholog 17 OrthoMCL 30 over representation 47 p value 47 50 PANTHER classification 16 PANTHER Generic Mapping File 43 49 PANTHER hit 38 PANTHER Pathway Applet 25 PANTHER protein class 10 12 PANTHER score 38 paralog 17 pathway 3 8 12 16 18 35 Pathway Accession 12 21 pathway detail page 21 pathway diagram 8 25 pathway home page 8 pathway list page 12 pathway molecule class 8 pathway molecule class detail page 22 Pathway Name 12 Pdeleterious 54 phylogenetic tree 7 28 pie chart 3 6 11 24 25 Process Description 26 protein class 3 16 35 Protein ID 9 16 prowler 2 34 PubMed 23 rank sum test 48 reference 21 22 reference gene list 44 RefSeq 6 registration 3 SBGN 26 SBML 8 sort 10 speciation 28 Species 35 Standard View 26 subfamily node 28 subPSEC 53 54 Supported ID 37 supported ID 42 synon
5. Name The name that appears on the pathway diagram It is usually an acronym or a short version of the full name Full name The complete more descriptive version of the name Synonyms All other names used to describe the component Definition A short description of the component Category ID Each component is considered as a pathway ontology term and is assigned with an ID It is usually a 5 digit number preceded by letter s to indicate the type of category e g G for DNA or RNA and P for protein Upstream A list of component s that modifies the current component in the pathway It usually implies that these components have protein protein interaction Downstream A list of component s that is modified by the current component in the pathway It usually implies that there are protein protein interaction involved Number Subfamilies Number of subfamilies that have training sequences directly as sociated with the component Clicking on the number will lead to the subfamily list page section 2 2 2 Number Associated Sequences Number of genes that are directly associated with the component Clicking on the number will lead gene list page section 2 2 1 GO classifications and PANTHER protein class The ontology terms classified to the genes that are associated to the pathway component Clicking on an ontology terms will lead to the Ontology Term Details page section 2 3 3 References A list of literature references or websites used to
6. PROTEIN Select upstream PiP2 SIMPLE MOLECULE Select upstream path PiP3 SIMPLE MOLECULE j mut pei bp A SHIP2 PROTEIN Select downstream pati j Tscl PROTEIN P Jj View Detail Tsc2 PROTEIN s Pathway Link p1l10alpha GENE p27 Apoptosis PHENOTYPE p53 PROTEIN g CH I2 i Figure 2 21 Pathway Diagram page 2 4 2 2 Export data from this page Users can export pathway related data using the following options CHAPTER 2 PANTHER AT A GLANCE 27 From the Export drop down menu all the pre generated images and SBML file of the pathway can be exported From the applet panel under File select Export image to export the current pathway diagram This function allows you to export the modified image 2 4 23 Using the pathway diagram The diagram page allows you to make selections on pathway components and then generate gene lists transcript lists or family lists You can use any of the following ways to select a single or multiple components 1 3 4 You can select a component by clicking on the name of the component in the left list panel of the applet If you want to select more than one component press and hold the control key while you select You can clear your selection by going to Select Clear all on the applet menu bar You can select components from the right diagram panel by clicking on the component symbol
7. and PANTHER Protein Classes and links to download them Section 2 1 4 Tools Links to all the PANTHER tools page Section 2 1 5 See details in Chapter 4 Workspace Links to the workspace available to the registered users Section 2 1 6 Registration is free Login Click here to login to the website and use the workspace PANTHER website does not require a login but the workspace is only available to registered users Registration It is not required to register to use the PANTHER website Registration allows you to create a workspace that you can store your analysis data on our site and it is free Help You can find more help tips at this link Whole genome function views Links to the whole genome pie chart Section 2 4 1 Gene expression tools Links directly to the Gene Expression Analysis Tools page Section 4 1 cSNP tools Links directly to the cSNP Scoring tool page Section 4 2 Upload multiple gene IDs Links directly to the Batch ID Upload page Section 3 3 Community Curation This page is currently provides the link that allows you to download PAINT Phylogenetic tree Annotation INference Tool application My Workspace Links to the workspace available to the registered users Section 2 1 6 Registration is free HMM scoring Links to the PANTHER HMM Scoring page Section 3 4 Downloads Links to the PANTHER FTP site Section 3 5 Genome statistics Links to page
8. lists all 165 PANTHER pathways and links to view the interactive pathway diagram download SBML and BioPAX files e Download pathway data it links to the ftp site for all pathway data download Section 3 5 lassification m Classificalion Sylo LOGIN REGISTER CONTACTUS HAP Community Pathway Curation Browse Pathways Search Pathways Pathway Resources PANTHER is giving a tutorial at the ICSB 2011 in Heidelberg on August 27 2011 Please click here for details Search PATHWAYS Pathways PANTHER Pathway consists of over 165 primarily signaling T pathways each with subfamilies and protein sequences Go mapped to individual pathway components A component is usually a single protein in a given organism but multiple Quick links proteins can sometimes play the same role Pathways are drawn using CellDesigner software capturing molecular level Whole genome function views Gene expression tools cSNP tools Upload multiple gene IDs Community Curation My Workspace HMM scoring Downloads Genome statistics Site map Newsletter subscription Enter your Email events in both signaling and metabolic pathways and can be exported in SBML format The SBGN view of the diagram can also be exported Pathway diagrams are interactive and include tools for visualizing gene expression data in the context of the diagrams Pathway release calendar NEW Community Pathway Curation Step by step i
9. not ding the reference g components only in the ren list Click on pathway name to see genes highlighted on pathway diagram eaS View Please select a chart to display m Homo sapiens genes REF samplel sample2 Pathways expected APvalue expected Pvalue FGF signaling pathway 124 60 37 4 57E 133 60 37 4 57E 133 EGF receptor signaling pathway 135 45 41 1 23E 84 35 41 5 46E 60 Unclassified 17337 0 2 24 491E 54 2 52 24 3 96E 49 Ras Pathway 79 21 24 2 55E 35 14 24 3 50E 21 Angiogenesis 191 24 58 9 53E 33 29 58 2 57E 42 Integrin signalling pathway 181 22 55 1 25E 29 17 55 5 28E 21 T cell activation 102 19 31 5 06bE 29 19 31 5 06E 29 VEGF signaling pathway T5 k FA 23 2 06bE 27 15 23 1 98bE 23 B cell activation 82 17 25 9246 27 21 25 5 55E 35 p53 pathway 113 18 34 2 75E 26 2 34 4 59E 02 Figure 4 6 Results of the binomial distribution analysis Tip Binomial distribution test Expected Value The expected value is the number of genes you would expect in your list for a particular PANTHER category based on the reference list As an example say we do do the analysis in terms of biological processes and we are interested in determining if there is over or under representation of chromatin packaging and remodeling genes in your uploaded list 4 1 1 4 CHAPTER 4 PANTHER TOOLS 47 As an example there are 23 481 genes in your reference list ex the human genome 1
10. 1mkllwalmkvlllallvslLagcqakmkglavllalalltgcqars mkv1l allv Lagcqak 4 4 44 4 1 q sequence Ll MKVLWAALLVTFLAGCQAKVEQAVETEPEPELR QOTE 37 lpladepkqrWEealdrFwdYlrelqtladdvqeelkssQiskELdtLie qrWE al rFwdYlr qt1l vqeel ssQ EL L sequence 38 WQSGQRWELALGRFWDYLRWVQTLSEQVQEELLSSQVTQELRALMD 83 DTMtELkaYkeeLeeqLtPvaeetrarlskelqalqarLgaDMeDakeRl TM ELkaYk eLeeqLtPvaeetrarlskelqa qarLgaDMeD Rl sequence 84 ETMKELKAYKSELEEQLTPVAEETRARLSKELQAAQARLGADMEDVCGRL 133 tqYrgElqamleOQnteevRaRvssylrKLrKRLlkDaeeLqkrlavYqag qYrgE qaml Q tee R R s lrKLrKRL1 Da LqkrlavYqag sequence 134 VOYRGEVOAMLGQSTEELRVRLASHLRKLRKRLLRDADDLOQKRLAVYQAG 183 aregaergvealRerlePlleqgqdraqaklgslaealksQakelkerle aregaerg at Rerl Pl eqg r a gsla sequence 184 AREGAERGLSAIRERLGPLVEQGRVR AATVGSLAGQ 219 sqaedlkerleataeelrgrLeeagerlrdrLdevreqieevkakvee l er at e lr r ee gtr rdrLdev eq evtak eet sequence 220 PLQERAQAWGERLRARMEEMGSRTRDRLDEVKEQVAEVRAKLEEqa 265 easaacceacacaecccacsasa FePLvedlqrQWagLvEKvQaavkpsaaetPas Toe RA 444 FePLved qrQWagLvEKvQaav saa P s sequence 266 qgirlqaeafqarlkswFEPLVEDMQRQWAGLVEKVQAAVGTSAAPVP S 314 dnh lt dnh sequence 315 DNH 317 Figure 3 7 The score result file from the PANTHER HMM score page 3 5 Download This is the PANTHER FTP site You can reach this page by clicking the Downloads link on the Quick links panel on the left side of the Home page You can download the foll
11. 2 8 Pathway list page You can sort the list convert the list to a different ID type and save the list as described in Gene List section 2 2 1 Pathway component list page A pathway component also called molecule class represents a group of homologous proteins across various organisms that participate in the same specific biochemical reactions within the pathway A pathway component is always associated with training sequences within the PANTHER libraries thus it is linked directly or indirectly to the PANTHER family subfamily statistical models This page lists pathway components with the following information Figure 2 9 e Component Accession The accession assigned to each pathway component It is usually a 5 digit number proceeded by a letter to indicate the type of the component in the pathway G for DNA or RNA P for protein Clicking it will lead to the Pathway molecule class detail page section 2 3 5 e Component Name The display name of the component Clicking it lead to the Pathway molecule class detail page section 2 3 5 e Type Indicates the type of the component in the pathway eg gene protein CHAPTER 2 PANTHER AT A GLANCE 14 e Upstream and Downstream These two columns list the pathway component s immedi ately upstream or downstream of the component Upstream is defined as the component that controls or modulates the reaction that the current component is involved in Down stream component s a
12. CANFAIENSEMBL GPVPDCKKAAASDQPSTSSEDREPCV1 QGRPKNGC I VKTGBLMACPT AKKLKKRNKPCPVCROPIOQMIVLTYFP SF9 MONDOIENSEMBL GFLPDCKKNKMNDQPSTSSEDREPCVIBOSRPKNGCIVRTGHLMACPTBAKKLKKRNKPCPVCROPIQMIVLTYPL GFVPDCKKIKATEQPSTSSEDREPCVIBOsRPKNGCILRTGMLMACPTBAKKLKKRNKPCPVCROPIOMIVLTYFS SF9 CHICKIENT DIVPGKRRLPSAIRALOGTEEROHRALBHMPKOAEKWLRTSMVM LYVBKKTEEEEOTLPGVOAAHTNDCTNLEWL SF9 XENTRIENSEMBL ENSXET GPVPDCKKSKLTSQPSTCSEAREPCVIBOTRPKNGCIVRTGHLMACYTBAKKLKKRNKPCPVCREPIQMIVLTYPS SF9 DANREIZFIN ZDB GENE 9 GVVPDCFPSPATTOPSTSSEEREPCVIBOSRPKNGCIVRTGHLMACYTBAKKLKNRNKLCPVCREPIQSVVLTYMS SF9 FUGRUIENSEMBL ENSTR SANEACASSIHVTEPASDPSARDPCLIBOSRPKNGCIVRTGHLISCY KLKKRNKLCPVCRMPIQSVILTYLS SF9 CIOINIENSEMBL ENSCING00000 SYVTDGRVTENNREPMEDPDPTGLCNI RNNPNDATMVNNGHOFCCP RLKRKGKSCPVCROPIMLVLRNYIV SF9 ORYSJIENTREZ 43433101IUniProtKB Q0D693 PLMEVRVRSEMAQMCSAMPQPRGSCC IMHOTOQVDSLLYRCGBMCTCPNGADOLKSSNRSCPICQSPIEDVVRAEMN Y x 2 4 Figure 2 24 The multiple sequence aligmment of PANTHER family PTHR10360 viewed in the tree viewer 3 1 3 1 1 Chapter 3 PANTHER Basics Keyword Search There are two types of keyword search pages the simple keyword search and advanced keyword search Simple keyword search The simple keyword search function can be found on the PANTHER home page as well as all the subject main page section 2 1 It allows users to do a quick search of the PANTHER database Tips about the search e The search term
13. ENSTRUGO0000008338 ENSEMBL ENSTRUPO0000020879 Fugu rubripes LDO DANRE ZFIN ZDB GENE 990415 153 UniProtKB Q561Z0 Danio rerio LDO CIOIN ENSEMBL ENSCINGO0000001188 ENSEMBL ENSCINP00000002267 Ciona intestinalis LDO ORYSJ ENTREZ 4343310 UniProtKB Q0D693 Oryza sativa LDO Figure 2 13 Ortholog table of the gene detail page 2 3 2 Family subfamily detail page This page provides details of a PANTHER family or subfamily model It contains 2 sections 1 Family subfamily information The pages for family and subfamily information are slightly different The family detail page contains the following information Figure 2 14 e Family The name of the family model Family ID is in parentheses e g PTHR10000 e Subfamilies The number of subfamilies under the family Clicking on the number will lead to the Subfamily List page section 2 2 2 e PANTHER Links Two links will lead to the Tree and MSA pages section 2 4 3 e GO molecular function biological process cellular component These are Gene On tology terms from PANTHER GO Slim describing the function of the gene product e PANTHER Protein Class This isa PANTHER Index term describing protein classes e PANTHER Pathway This links to a diagram of the pathway s in which a gene product participates e Genes The number of genes that were used as training sequences to build the family model Clicking on the number will lead to the Gene List Page section 2 2 1 e HMM Length The number
14. You can clear your selection by clicking on the selected item again You don t need to hold the control key for multiple selections Under Select applet menu bar you can do the following e Select all proteins genes and RNA Since only proteins genes and RNA in PAN THER pathway are associated with PANTHER family genes you have an option to select all of them by one click e Select all This function allows you to select all items including simple molecules ions phenotypes etc e Clear all You can use this function to clear all your selections By right clicking on the component you can make the following selections e Upstream or Downstream component This function will select the immediate or downstream component If the selected component appears in multiple places in the same pathway they will be highlighted also e Upstream or Downstream path This function will select all components in the entire upstream or downstream path Once the selections are made you can generate any of the following lists by making the selection to the Search selection for drop down menu on the right side of the page Genes A list if genes whose proteins hit PANTHER family and subfamily models with at least one training sequence associated with the selected pathway component Transcripts proteins A list of proteins that hit PANTHER family and subfamily models with at least one training sequence associated with the selected pathway componen
15. components may have multiple genes mapped to them The color ranges for these components are determined by the following 9 median of the gene values default O average of the gene values O maximum gene value O minimum gene value Figure 4 13 Color code for the head map 4 2 Evolutionary Analysis of Coding SNPs This tool estimates the likelihood of a particular nonsynonymous amino acid changing coding SNP to cause a functional impact on the protein It calculates the subPSEC substitution position specific evolutionary conservation score based on an alignment of evolutionarily related proteins as described in Brunham et al 2005 9 Thomas et al 2003 10 and Thomas and Kejariwal 2004 11 4 2 1 Input protein and substitution data Paste the wildtype protein sequence in the first box FASTA format or just the protein sequence itself and then enter the substitution s relative to this input sequence in the standard amino acid substitution format e g A265V Multiple substitutions should be separated by a tab space or return Figure 4 14 4 2 2 4 2 2 1 4 2 2 2 4 2 2 3 CHAPTER 4 PANTHER TOOLS 54 EVOLUTIONARY ANALYSIS OF CODING SNPS Estimates the likelihood of a particular nonsynonymous amino acid changing coding SNP to cause a functional impact on the protein It calculates the subPSEC substitution position specific evolutionary conservation score based on an alignment of evolutionarily related pr
16. of position in the hidden Markov model e Downloads You can download the family HMM built with either HMMER or SAM method CHAPTER 2 PANTHER AT A GLANCE PANTHER FAMILY INFORMATION Family Subfamilies PANTHER Links GO Molecular Function GO Biological Process GO Cellular Component PANTHER protein class Pathway Categories Genes HMM Length Downloads Figure 2 14 Family MDM2 4 PTHR10360 Bw Tree MSA catalytic activity 5 ligase activity 4 ubiquitin protein ligase activit apoptosis 4 negative requlation of apoptosis ligase 4 ubiquitin protein ligase p53 pathway feedback loops 2 4 MDM 2 p53 pathway gt Mdm2 Insulin IGF pathway protein kinase B signaling cascade 4 Mdm2 p53 pathway 4 Mouse double minute 2 P53 pathway feedback loops 1 5 Mdm2 Insulin IGF pathway protein kinase B signaling cascade 4 Mdm2 P53 pathway feedback loops 1 4 Mouse double minute 2 p53 pathway feedback loops 2 G MDM 2 31 501 HMM HMMER format information section of the family detail page The subfamily detail page contains similar information as the family detail page except for the first two items Figure 2 15 e Subfamily The name of the subfamily and the subfamily ID eg PTHR10000 SF1 e Family The family name that the subfamily belongs to Clicking the name will lead you to the family detail page CHAPTER 2 PANTHER AT A GLANCE 20 PANTHER SUBFAMIL
17. ranked from smallest to largest with the smallest value getting a rank of 1 If multiple values are identical the average of the ranks for these values is used Then the rank sum is calculated for this category by summing up the ranks for all of the genes that map to this category The average rank R1 is then calculated by dividing the rank sum by the number of genes n1 that map to the category Likewise the rank sum is calculated for the list of all IDs uploaded and the average rank R2 is calculated by dividing the rank sum by the total number of genes uploaded n2 Next the Mann Whitney U statistic is calculated for both populations U1 nl n2 n1 nl 1 2 R1 U2 n2 n2 nl n2 1 2 R2 The larger of these two values is the Mann Whitney U statistic U whose distribution for small sample sizes can be found in most statistic books In our case our application is for large sample sizes so we use the normal approximation Z score U n1 n2 2 sqrt n1 n2 n14 n24 1 12 It follows that the p value is the integral under the standard normal density View results in pathway diagram If you select to analyze the data in PANTHER pathway you can click the pathway name from the column 1 of the result page and view the results in pathway diagram in head map Figure 4 12 The color range reflects the expression value of your genes mapped to the pathway compo nent You can define the color range by selectin
18. with statistics of 48 genomes in the PANTHER library Site map Links to the site map Keyword search Perform simple keywords search for genes proteins families ontology and pathway terms Section 3 1 1 Sequence search Enter a protein sequence to score against PANTHER HMM library Section 3 4 Newsletters The most recent newsletter is displayed All archived newsletters can be retrieved by clicking the additional info link CHAPTER 1 WELCOME TO PANTHER SYSTEM 4 24 Publications This section lists some of the PANTHER publications and information about citing PANTHER Chapter 2 PANTHER At A Glance PANTHER data are stored in a database in the backend PANTHER website presents the data to the users through a number of text pages and graph diagram pages The text pages include e Subject main page e List pages e Information detail pages The graph and diagram pages include e Pie charts e Pathway diagrams e Tree Viewer CHAPTER 2 PANTHER AT A GLANCE 6 2 1 Subject main page 2 1 1 Genes and orthologs PANTHER now contains the complete sets of protein coding genes for 48 different organisms obtained from various sources including Model Organism Database UniProt RefSeq and EN SEMBL This page serves as a portal for you to access information related to genes genomes and orthologs Figure 2 1 There are 4 links on this page that allows you to e perform simple keyword search Section 3 1 2 e fin
19. 000156510 UniProtKB Q2TB90 Putative Q2TB90 SUBFAMILY NOT kinase activity carbohydrate metabolic process carbohydrate kinase Glycolysis gt Hexokinase Homo sapiens hexokinase NAMED HKDC1 PTHR19443 SF0 Pentose phosphate HKDC1 pathway gt Hexokinase Fructose galactose metabolism gt Hexokinase O 2 DROME FB FBgn0042711 UniProtKB Q9NFT9 Hexokinase Q9NFT9 SUBFAMILY NOT kinase activity carbohydrate metabolic process carbohydrate kinase Glycolysis gt Hexokinase Drosophila type 1 NAMED melanogaster Hex ti PTHR19443 SFO Pentose phosphate pathway gt Hexokinase Fructose galactose metabolism gt Hexokinase 3 HUMAN ENSEMBL ENSGO0000156515 UniProtKB P19367 Hexokinase 1 P19367 SUBFAMILY NOT kinase activity carbohydrate metabolic process carbohydrate kinase Glycolysis Hexokinase Homo sapiens HKI NAMED PTHR19443 SFO Pentose phosphate pathway gt Hexokinase Fructose galactose metabolism gt Hexokinase O 4 MOUSE MGI MGI 96103 UniProtKB P17710 Hexokinase 1 P17710 SUBFAMILY NOT kinase activity carbohydrate metabolic process carbohydrate kinase Glycolysis gt Hexokinase Mus musculus Hki NAMED PTHR19443 SF00 Pentose phosphate pathway gt Hexokinase Fructose galactose metabolism gt Hexokinase 5 RATIRGD 2796 UniProtKB P05708 Hexokinase 1 P05708 SUBFAMILY NOT kinase activity carbohydrate metabolic process carbohydrate kinase Glycolysis gt Hexokinase Rattus norvegicus Hkl NAMED PTHR19443 SF0 Pentose phosphate pathway Hexokinase Fructose ga
20. 000691 PC00142 lig CHICKIENT MDM4 P53 BIND PREDICTED hypot Gallus galus ENTREZ 41 MDM4 GO 001687 GO 000691 PC00142 lig XENTRIEN MDM4 P53 BIND Protein Mdm4 p53 b Xenopus tro ENSEMBLE GO 001687 GO 000691 PC00142 lig DANREIZFI MDM4 P53 BIND Protein Mdm4 Danio rerio ZFIN ZDB mdm4 GO 001687 GO 000691 PC00142 lig FUGRUIEN MDM4 P53 BIND Protein Mdm4 p53 b Fugu rubripes ENSEMBLE GO 001687 GO 000691 PC00142 lig PANTRIEN E3 UBIQUITIN P E3 ubiquitin protein li Pan troglody ENSEMBLE GO 001687 GO 000691 PCO00142 lig HUMANIEN E3 UBIQUITIN P E3 ubiquitin protein li Homo sapiens ENSEMBL E MDM2 GO 001687 GO 000691 PCO00142 li MACMUIEN E3 UBIQUITIN P E3 ubiquitin protein fi Macaca mul ENSEMBLE GO 001687 GO 000691 PCO00142 lig RATIRGD E3 UBIQUITIN P PREDICTED similar Rattus norv RGD 1305332 GO 001687 GO 000691 PCO00142 li MOUSEIMG E3 UBIQUITIN P E3 ubiquitin protein fi Mus musculus MGI MGI 9 Mdm2 GO 001687 GO 000691 PC00142 lig BOVINIENS E3 UBIQUITIN P MDM2 protein Bos taurus ENSEMBL E MDM2 GO 001687 GO 000691 PCO00142 lig BOVINIENS E3 UBIQUITIN P Bos taurus ENSEMBLE GO 001687 GO 000691 PCO00142 lig CANFAIEN E3 UBIQUITIN P E3 ubiquitin protein li Canis familia ENSEMBL E MDM2 GO 001687 GO 000691 PCO00142 lig MOND
21. 14 P00905 PTEN Protein PTEN Insulin IGF pathway protein kinase B signaling cascade 18 Z Q 15 p02798 cyt C Protein ATP synthesis 40 2 O 16 P04494 Tsc2 Protein Protein kinase B Insulin IGF pathway protein kinase B signaling cascade 5 T O 17 P04495 Isci Protein Protein kinase B Insulin IGF pathway protein kinase B signaling cascade 5 1 O 18 P04496 Mdm2 Protein Mdm2 p53 Insulin IGF pathway protein kinase B signaling cascade 11 32 Protein kinase B 19 P04497 253 Protein Mdm2 piiOalpha Insulin IGF pathway protein kinase B signaling cascade 0 0 Mdm2 PTEN 20 P04498 Pil0alpha Protein pliOalpha Insulin IGF pathway protein kinase B signaling cascade 10 Figure 2 9 Pathway component list page You can sort the list convert the list to a different ID type and save the list as described in Gene List section 2 2 1 2 2 5 Ontology term list page This page provides a list of PANTHER ontology terms including PANTHER GO Slim and PANTHER Protein Class with the following related information e Category ID A GO id for PANTHER GO Slim or a PANTHER Protein Class id which is a 5 digit number preceded by letters PC Clicking on the ID will lead to the Ontology Term Detail page section 2 3 3 2 3 2 3 1 CHAPTER 2 PANTHER AT A GLANCE e Name Category names 15 e Parent The names of all parent categories A parent category refers to a more general category of the current one e g Receptor is a parent category o
22. 24 Edit Delete Move Share 2011 08 24 22 03 O ES uniProtFly 7035 Edit Delete Move Share 2008 01 31 18 19 All Uniprot fly candidateGene 3 20 2 Edit Delete Move Share 2006 03 23 14 10 The final candi 2 dme gene info for Til Edit Delete Move Share 2005 05 11 00 00 Figure 3 8 Screenshot of a sample workspace page CHAPTER 3 PANTHER BASICS Al To access your Workspace you just need to click the Workspace link at the PANTHER Home page Section Jrefsec ov and login You will then reach the main Workspace page Figure 3 8 The exported list is in the root folder by default You can organize the lists by using the Move link in the second column of the table to move the list to the appropriate folder You can also create a new folder by clicking the create new folder icon on the top left part of the table Here are some other things you can do to your lists e Go to the list by clicking the list name in the first column The number in the parenthesis indicates the number of entries e g genes in the list Share the list with other registered PANTHER users by clicking the Share link or the icon You then need to enter the username of the Workspace that you would like to share Modify the name and description by clicking the Edit button Delete the list 4 1 4 1 1 4 1 1 1 Chapter 4 PANTHER Tools In this chapter we will describe the PANTHER tools for experime
23. 25 of these genes map to the Chromatin packaging and remodeling category Based on this 0 532 125 divided by 23481 of the genes in the reference list are involved in chromatin packaging and remodeling Now your upload list contains 978 genes Based on the reference list we would expect that 5 21 genes 978 0 532 in your uploaded list would be involved in chromatin packaging and remodeling If for this biological process you observe more genes in your uploaded list than expected you have an over representation of genes involved in chromatin packaging and remodeling If you observe fewer genes than expected you have an under representation P Value calculated by the Binomial statistic In the example above we expected 5 21 genes in your uploaded list to be involved in chro matin packaging and remodeling Say that we actually observe 7 genes involved in this biological process This is very similar to 5 21 so you would expect a large p value the p value would be 0 27 Alternatively if 15 of your uploaded genes map to this process this is very different than the expected p value so you would expect a small significant p value the p value would be 3 3E 04 This small p value indicates that the result is non random and potentially interesting and worth looking at in closer detail A p value cutoff of 0 05 is recommended as a start point The binomial statistic is a commonly used statistic it can be found in any statistics boo
24. 6 UniProtK B ID UniProt ID Example AGAP3 HUMAN e PANTHER Generic Mapping File list type If your id type is not listed in the above supported ID list select this list type and you can analyze IDs not in the PANTHER system by uploading user generated data containing mappings between any arbitrary identifier and its corresponding PANTHER ID see below for details about mapping You can map your sequences to PANTHER by downloading the PANTHER HMM scoring tools available at the downloads section on the PANTHER site and scoring your sequences against the PANTHER HMM library For sequences that do not match a PANTHER HMM please assign a PANTHER ID of NOHIT This is very important since this information will be used to assign sequences to the unclassified category This will ensure that the statistics can be calculated correctly e File format The tool only accepts simple text txt in the following format If your file contains a list of IDs supported by PANTHER separate the IDs by tab carriage return line return comma pipe or space If you are uploading PANTHER Generic Mapping File the file must be tab delimited and contain the following columns The first column can contain any arbitrary ID ex a probe ID but must be unique since this allows the user to uniquely specify each record in the dataset so they can track the identifier on the PANTHER website The second column should be corresponding PANTHER I
25. D ex PTHR10078 or PTHR10078 SF1 and is used to look up the molecular function biological process and pathway associations Microsoft Excel file format is not accepted by the tool To convert an Excel format xls or xlsx to a simple text format txt Open the file in Excel under File menu choose Save As and under the Format option choose Tab Delimited Text and then click the Save button 2 Choose from your Workspace Figure 4 3 If you registered and have a Workspace you can upload lists that saved in the Workspace directly for analysis See Workspace section for its details CHAPTER 4 PANTHER TOOLS 44 Steps 1 Select Lists to Compare to a Reference List 1 Select list s to For example each selected list may be a cluster of co expressed genes under a particular analyze set of conditions Figure 4 1 Select a list to analyze UPLOAD OR SELECT LIST FROM YOUR WORKSPACE Uploaded and selected lists Upload list ust ype Please upload a fio or select a ist Upload list I Fished selecting nts If there are redundant IDs only the first will be used in the analysis Please login to select lists from your workspace Figure 4 2 Upload a gene list Upload list Uploaded and selected lists List type 3 Please select list type E Please upload a file or select a list Upload list RIDES If there are redundant IDs only the first wil be used in the analysis Choose list from your workspace
26. Diagram The Activity Flow view is the default view of the pathways You can toggle the two view by clicking the tab on the top of the applet window e SBGN Image View When this tab is selected a static non interactive image of SBGN Process Description Diagram is displayed INSULIN IGF PATHWAY PROTEIN KINASE B SIGNALING CASCADE M BGN created vt Export Select one E L CellDesigner Click components to make selections Right click components for more options Search selections for Pathway Diagram Genes Genes File Edit Zoom Select Color Transcripts Proteins Model Interactive Standard Activity Flow iew g PANTHER Categories m i Sncinragelriees PANTHER Families v Molecules Pathways 5 ADP SIMPLE MOLECULE Standard View Activity Flow B pathway Components ATP SIMPLE MOLECULE Cell growth low ROS high HSP PHENC Cell survival maintenance PHENOTYF Complex Insulin IGF INSR IGF R IRR Complex PI3 K P110alpha COMPL Complex Tsc1 Tsc2 COMPLEX Complex Tsc2 Tsc1 COMPLEX FKHR PROTEIN GSK 3 PROTEIN Glucose depreivation response PHEN Glycogen synthesis PHENOTYPE INSR IGF R IRR PROTEIN IRS 1 4 PROTEIN Insulin IGF PROTEIN Mdm2 GENE P110alpha PROTEIN P53 pathway PHENOTYPE PDK1 2 PROTEIN PI3 K PROTEIN PKB PROTEIN PTEN GENE 2 PTEN
27. OIEN E3 UBIQUITIN P E3 ubiquitin protein li Monodelphis ENSEMBLE GO 001687 GO 000691 PCO00142 lig ORNANIEN E3 UBIQUITIN P E3 ubiquitin protein li Ornithorhyn ENSEMBLE GO 001687 GO 000691 PCO00142 lig CHICKIENT E3 UBIQUITIN P PREDICTED similar Gallus galus ENTREZ 39 MDM2 GO 001687 GO 000691 PC00142 lig XENTRIEN E3 UBIQUITIN P Mdm2 p53 binding pr Xenopus tro ENSEMBL E mdm2 GO 001687 GO 000691 PC00142 lig DANREIZFI E3 UBIQUITIN P Mdm2 protein Danio rerio ZFIN ZDB mdm2 GO 001687 GO 000691 PCO00142 lig FUGRUIEN E3 UBIQUITIN P E3 ubiquitin protein li Fugu rubripes ENSEMBLE GO 001687 GO 000691 PC00142 lig I CIOINIENS E3 UBIQUITIN P Ciona intesti ENSEMBLE GO 001687 GO 000691 PC00142 lig LORYSJIEN BIQUITIN P B i a sativa 4 20499 00168 00069 X io LY Figure 2 23 A tree viewer with expanded attribute table 2 4 3 3 MSA When the MSA button is clicked on the right panel the multiple sequence alignment view is shown You can toggle back to the attribute table view by clicking the Grid button PANTHER multiple sequence alignments MSAs are the basis for the PANTHER distance trees and therefore of the family subfamily classification MSA is built using the MAFFT software 5 MSA is aligned across the entire length of all sequences in the family The evolutio
28. PANTHER User Manual For PANTHER 7 0 Date September 19 2011 Authors The PANTHER Team Contents 1 Welcome to PANTHER System 1 1 About this document 1 2 How to cite PANTHER 1 9 PANTHER W W 2 sn RR WE OWOIVIGW usu imum lm e Soe we PANTHER At A Glance 2 1 Subject main page 2 1 1 Genes and orthologs 2 1 2 Trees and HMMs 2 1 3 Pathways zd QhOogies ckc6x iex 2 1 5 00lS 4 wis GOA an RERO 2 1 6 Workspace 22 LSRpages c9 FR aw eS a ee 4 2 2 1 Genelistpage 2 2 2 Family subfamily list page 2 2 3 Pathway list page 2 2 4 Pathway component list page 2 2 5 Ontology term list page 2 3 Information detailpages 2 8 4 Gene detail page 2 3 2 Family subfamily detail page 2 3 3 Ontology term detail page 2 3 4 Pathway description page N Ln XQ dq dq DOAN DOA 2 3 5 Pathway molecular class compo nent detailpage 2 4 Graph and diagram pages 241 Piecharts ci e 2 bw cae FAS 2 4 2 Pathway diagram 2 4 8 TreeViewer PANTHER Basics 3 1 KeywordSearch 3 1 1 Simple keyword search 3 1 2 Advanced keyword search 9 2 Prowlef 25229364 3 Batch ID Search 2 omo Ros 3 4 PANTHER HMM Score 35 Download 2c 04 RA emu 3 6 Workspace PANTHER Tools 4 1 Gene expression data analysis tools 4 1 1 Compare Gene List 4 1 2 Analyze gene list with expression VaIUGS 2 m s 4 2 Evolutionary Analysis of Coding SNPs 4 2 1 Inpu
29. PANTRIENSEM PANTRIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Pan troglody ETE SF9 HUMANIENSE HUMANIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Homo sapiens EN f MACMUIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Macaca mul SF9 RATIRGD 13 RATIRGDs E3 UBIQUITIN PROTEIN LIGASE MDM PREDICTED similar to trans Rattus norv R SF9 MOUSEIMGI MOUSEIMG E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Mus musculus M SF9 BOVINIENSEME BOVINIENS E3 UBIQUITIN PROTEIN LIGASE MDM MDM2 protein Bos taurus BOVINIENS E3 UBIQUITIN PROTEIN LIGASE MDM Bos taurus SF9 CANFAIENSEMI CANFAIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Canis familia SF9 MONDOIENSEMI MONDOIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Monodelphis ORNANIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Ornithorhyn SF9 CHICKIEI CHICKIENT E3 UBIQUITIN PROTEIN LIGASE MDM PREDICTED similar to MDM2 Gallus gallus SF9 XENTRIENSEMBL ENSXI XENTRIEN E3 UBIQUITIN PROTEIN LIGASE MDM Mdm2 p53 binding protein ho Xenopus tro SF9 DANREIZFIN ZDB GENE DANREIZFI E3 UBIQUITIN PROTEIN LIGASE MDM Mdm2 protein Danio rerio SF9 FUGRUIENSEMBL ENS FUGRUIEN E3 UBIQUITIN PROTEIN LIGASE MDM E3 ubiquitin protein ligase Md Fugu rubripes EN SF9 CIOINIENSEMBL EN
30. PHGSESTEQEKPCSL EKRPRDGNIIKTSBLTTCFH LKKSGASCPACKKEIQLVIKVEI SF10 MOUSEIMC GIVPDCRRTISAPHSSESAEQEKPCSL EKRPRDGNIIKTSBLTTCFH KKSGASCPVCKKEIQLVIKVPI SF10 BOV GVVPDCRRTVSAPHSSESMEQEKPCSLBEKRPRNGNIIRTGMLVTCFEH LKKAGASCPICKKEIQLVIKVEV SF10 BOV GIVPDCRRTVSAPHSSESMEQEKPCNLBEK1PONGNIIRTGLVTCEH LKKAGASCPICKKEIQLVIKVEV SF10 MONDOIENSEMBL t GIVPDCRRTISDPQSSESLEQEKPCKLBEKRPRDGNIIRTAMLVVCE RLKKTOAPCPICKKSIQMVIKIEV SF10 ORNANIENSEMBL EN GIVPDCRRTISAPEGSESLEQEELCSLBEKRLRDGNIVRTAMLVACE KLKKGGAPCPVCKKDIQMVIKTEVV SF10 CHICKIENTREZ 41982 GIVPDCRRTISAPRECESFEHKNPCPLBHHRPRDGNIVRTAMLVACE MLKKRRSPCPVCRKDIEMVIRIPMG SF10 XENTRIENSEMBL ENS GLMPDCRRTVSAPANSKALKSTEPCRL OQRRORNGNVVRT VTCFS NLKKNRKGCPVCOKPIOMVVKIYV SF10 DANREIZFINZZDB GENE GIMPDCLRTVSDPSSSKQIHHEEPCKL RVRPRNGNIIRT ITCFP KLHKFHAPCPGCGQVIQKVIKTFI SF10 FUGRUIENSEMBL ENST GIVPDCTRTVSDPLSGKSSHKEEPCKLBRVRPRNGNIIRTAMLLTCFPBARRLBKPHAPCPGCGKIIQNVIKIPIL SFS PANTRIENSEMBL EPCVIBOGRPKNGCIVKTGHLMACPTBAKKLKKRNKPCPVCROPIOMIVLTYFPP SF9 HUMANIENSEMB GPVPDCKKT IVNDQPSTSSEDREPCV1G QGRPKNGC I VKTGBLMACPT AKKLKKRNKPCPVCROPIOQMIVLTYFP GPVPDCKKTIVNDQPSTSSEDREPCVIBOoGRPKNGCIVKTGHLMACPTBAKKLKKRNKPCPVCROPIOMIVLTYFP SF9 RATIRGD 13053 GLVPDGKKVTEDDQPSTSSESKEPCV1IG QGRPKNGC I VKTGBLMSCPT AKKLKKRNKPCPVCROPIOQMIVLTYFN SF9 MOUSEIMGI MC GLVPDGKKLTENDQPSTSSESKEPCVIBOGRPKNGCIVKTGHLMSCPTBAKKLKKRNKPCPVCROPIQMIVLSYEN SF9 BOVINIENSEMBL GFVPDCKKSTVSDQOPSTSSEDREPCVIBOGRPKNGCIVKTGHLMACPTBAKKLKKRNKPCPVCROPIQMIVLTYPFP SF9
31. R Classification System ll You may select a combination of Biological Processes Molecular Functions Pathways or Species of interest by clicking on the checkbox next to each item s name Il Once you have finished making your selections you may choose which type of result you re interested in from If the Prowler is not loading correctly click here Figure 3 4 The PANTHER Prowler e Ontology Type red arrow On the left side of the Browser is the Ontology Type panel There are five ontology types you can choose Biological Process this is a GO Slim biological process ontology Molecular Function this is GO Slim molecular function ontology CHAPTER 3 PANTHER BASICS 35 Cellular Component this is a GO Slim cellular component ontology Protein Class this is PANTHER ontology to categorize protein families Pathway The last choice is species You can choose the species when querying over certain result sets such as genes to narrow your search There are 48 total species that you can choose with 12 model organism species listed first and the 36 remaining in alphabetical order The default is all species e Ontology blue arrow This panel is in the middle of the prowler and it changes based on the selection of the ontology type You can click on the to expand the next level of the ontology terms The details of each ontology term can be viewed by mouse over the name or by clicking th
32. SCING0Od CIOINIENS E3 UBIQUITIN PROTEIN LIGASE MDM Ciona intesti SF9 ORYSJIENTREZ 4343310lUniProtKB Q0D693 ORYSJIEN E3 UBIQUITIN PROTEIN LIGASE MDM Os0790499800 protein Oryzasativa El EK gt G PANTHER Classification System Figure 2 22 A phylogenetic tree of PANTHER family PTHR10360 viewed in the tree viewer 2 4 3 2 Attribute table The attribute table contains one row for each sequence in the tree Each column displays a different attribute of the sequences as follows Figure 2 23 e Protein id This is the long id for each sequence The format of the id is as below organism gene DB gene id protein DB protein id When the tree is collapsed this col umn shows the annotation node AN ids e sf name The name annotated for the subfamily e definition This is the brief definition line parsed out from the protein database where the protein sequence is from CHAPTER 2 PANTHER AT A GLANCE 30 e organism This is the organism from which the sequence was derived Click on the organism name to open the full taxonomy record for that organism e gene id e gene symbol e OrthoMCL The OrthoMCL id that the gene is predicted to belong to e Molecular function Biological Process and Cellular component These are Gene Ontology terms from PANTHER GO Slim describing the function of the gene product e Protein Class This is a PANTHER Index term de
33. Y INFORMATION Subfamily E3 UBIQUITIN PROTEIN LIGASE MDM2 P53 BINDING PROTEIN MDM2 PTHR10360 SF9 Family MDM2 4 PANTHER Links amp IE SA Tree GO Molecular Function catalytic activity 4 ligase activity 4 ubiquitin protein ligase activity Figure 2 15 Subfamily information section of the family detail page 2 Genes Assigned To This Family A table listing the number of genes from various species that hit this model Clicking on the number will lead to the Gene List page section 2 2 1 GENES ASSIGNED TO THIS FAMILY Species Count Bos taurus Canis familiaris Ciona intestinalis Danio rerio Fugu rubripes Gallus gallus Homo sapiens Macaca mulatta Monodelphis domestica Mus musculus Ornithorhynchus anatinus Oryza sativa Pan troglodytes Rattus norvegicus IN IN N de N N N WU N N N Ni d Xenopus tropicalis Figure 2 16 List of genes assigned to the family in the family detail page 2 3 3 Ontology term detail page This page provides the following information for a particular PANTHER GO Slim term or a PANTHER protein class category Figure 2 17 Name The name of the category Definition brief description of the category Class ID A GO id for PANTHER GO Slim or an internal tracking ID for PANTHER Protein Class The PANTHER id is usually a 5 digit number preceded by letters PC for Protein Class Parent lineages The path of the category with all parent categories For GO te
34. access information related to the trees and HMMs Figure 2 2 There are 4 links on this page that allows you to e search for PANTHER families trees and HMMs This basically takes you to the keyword search page Section 3 1 2 e browse families and subfamilies by classifications which takes you to the prowler page Section 3 2 e score a new sequence against PANTHER HMMs which takes you to the HMM score page Section 3 4 e download the PANTHER scoring tool to score multiple sequences against PANTHER HMMs Section 3 5 lassification m ACO SUUM LOGIN REGISTER CONTACTUS HEAP Home Browse CDITEDPBENUPIPHE Trees and HMMs Ontologies Tools Workspace Search Families Download HMMs PANTHER is giving a tutorial at the ICSB 2011 in Heidelberg on August 27 2011 Please click here for details Search GENE FAMILIES PHYLOGENETIC TREES HMMS PANTHER version 7 0 contains 6594 protein families each with a phylogenetic tree relating modern day genes in 48 organisms Go Phylogenetic trees also infer the evolutionary events that occurred namely speciation green nodes and gene duplication Quick links events orange nodes Horizontal transfer events are not inferred in this version of PANTHER but will appear in future Whole genome function versions views Expert biologists have divided each family into subfamilies which are generally orthologous groups but may also contain cSNP tools recently duplicated paralogs Each fa
35. ative requlation of apoptosis GO Cellular Component PANTHER protein class ligase 4 ubiquitin protein ligase Pathway Categories p53 pathway feedback loops 2 b MDM 2 Ubiquitin proteasome pathway 4 Ubiquitin protein ligase E3 p53 pathway 5 Mdm2 Insulin IGF pathway protein kinase B signaling cascade L Mdm2 p53 pathway 4 Mouse double minute 2 P53 pathway feedback loops 1 5 Mdm2 Insulin IGF pathway protein kinase B signaling cascade L Mdm2 P53 pathway feedback loops 1 4 Mouse double minute 2 p53 pathway feedback loops 2 4 MDM 2 Figure 2 12 Gene classification section of the gene detail page 3 Orthologs Figure 2 13 This section lists all the orthologs of this gene e LDO means least diverged ortholog while other more diverged orthologs if any are marked O Paralogs are marked P e For orthologs the species column gives the modern day species in which the ortholog is found For paralogs the species column gives the two speciation events between which the duplication occurred that generated the paralogous genes ND means not determined Thus different paralogs can be distinguished by how long ago the relevant duplications occurred e Orthologs are genes that can be traced to the same gene in the genome of their most recent common ancestor species Paralogs are genes that are traced to related but distinct genes in the genome of their most recent common ancestor
36. ber of components or molecule classes in each pathway Clicking the number will take you to the pathway component list page e Subfamilies This column provides the number of subfamilies of all genes that are as sociated to each pathway Clicking the number will take you to the Subfamily list page section 2 2 2 e Associated Sequences This column lists the numbers of genes that are associated to the pathway Clicking the number will take you to the Gene list page section 2 2 1 PANTHER PATHWAY LIST Convert List to Select Send list to Select HJ Display 30 ie Items per page Refine Search Hits 1 30 of 33 page 1 2 er an A Pathway Pathway Name Components Subfamilies Associated Accession Sequence Q 1 Po0005 Angiogenesis ee 202 1258 D 2 P00008 Axon quidance mediated by Slit Robo 14 rx AzL o 3 P00010 B cell activation 37 54 455 g 4 P00012 Cadherin signaling pathway 16 153 1050 p 5 P00016 Cytoskeletal regulation by Rho GTPase 20 66 807 j 6 P00018 EGF receptor signaling pathway 28 176 1181 D 7 P00021 FGF signaling pathway 26 124 1149 O 8 P00023 General transcription requlation 19 63 379 p 9 P00029 Huntington disease 60 184 1528 10 P00030 Hypoxia response via HIF activation 14 37 200 11 P00031 Inflammation mediated by chemokine and cytokine 58 249 1638 signaling pathway Q 12 P00033 Insulin IGF pathway protein kinase B signaling 18 52 275 cascade Figure
37. can be an identifier a word or a phrase multiple terms e The search looks for the exact match of the entire word For example if you enter the term hexo the search will not find hexokinase Therefore if you are not sure about the word a wild card can be used In this case if you enter hexo the search will find hexokinase e You can specify a particular subject to search from the drop down menu such as Genes and orthologs or Pathways The default on the home page is All and the default on each subject main page is the subject itself e The search looks for the search term s in all fields of the selected subject Some of these fields may not be visible to you on the result list page such as synonyms definitions KEYWORD SEARCH Genes and orthologs Pathways Families Ontology terms Figure 3 1 Simple keyword search 32 3 1 2 CHAPTER 3 PANTHER BASICS 33 Advanced keyword search The result of the simple keyword search will lead to the advanced keyword search page where you can refine your search or you can display the results list pages This search page can also be accessed from the keyword search link in the Genes and orthologs main page section 2 1 1 Enter keyword s mdm2 EJ Help SEARCH RESULTS 3 Genes E Families at Pathways E Ontology terms SELECTED GENOMES Select species for gene search Close M Homo sapiens V Mus musculus M Rattus norv
38. cation file Contains the PANTHER family subfamily GO slim PANTHER protein class and pathway classifications for the the complete genomes derived from the 48 organisms used in the PANTER database e PANTHER HMM Classification file Contains the PANTHER family subfamily name and the GO slim PANTHER protein class and pathway classifications for every PAN THER protein family and subfamily in PANTHER HMM library e PAINT PAINT is a java application for viewing tree files Minimum Java version 1 4 required To install uncompress the download file and execute launchTAV bat for windows or sh launchTAV sh for MAC 3 6 Workspace If you have registered and have a Workspace account in PANTHER here is a description about how you can use it You can export any list from the list page to the Workspace Typically after a keyword search prowler search or batch search you would like to save the resulting list so that you do not have to perform such search over and over Workspace allows you to do so When you use the export to workspace function on any of the list page see description is section 2 2 1 you will be asked to provide a name required and a description optional to the list my workspace oF Options Date Owner Description Root Folder N A C 2 sample 2 50 Edit Delete Move Share 2011 08 24 22 06 C sample 1 50 Edit Delete Move Share 2011 08 24 22 05 C sample 1
39. ch is the number of genes you would expect in your list for this PANTHER category based on the reference list See below for more detailed explanation The fifth column has either a or A plus sign indicates over representation of this cat egory in your experiment you observed more genes than expected based on the reference list for this category the number of genes in your list is greater than the expected value Conversely a negative sign indicates under representation The sixth column is the p value as determined by the binomial statistic This is the probability that the number of genes you observed in this category occurred by chance randomly as determined by your reference list A small p value indicates that the num ber you observed is significant and potentially interesting A cutoff of 0 05 is recommended as a starting point See below for more explanation If you upload more than one list the 3 6 columns are repeated for each list The result is sorted based on p values of the first list but you can click on any column header to sort based on that column The yellow triangle in front of the header indicates which column is sorted and the orientation of the triangle indicates whether the sort is ascending or descending Results Reference list samplel sample2 In pathway diagr Mapped IDs 19911 60 60 samplel red Unmapped IDs 0 0 0 sample2 green yellow components in ist
40. column and view pathway diagram with your IDs overlay on the diagram Each list has an assigned color as shown on the top of the result page Figure 4 6 The drop down menu allows you to select a color of choice for each list CHAPTER 4 PANTHER TOOLS 48 Figure 4 7 shows an example of such a diagram Red and green indicate the genes appearing in each of your list Yellow indicates that the pathway component is in more than one of the uploaded list not including the reference list Gray indicates the components not in your list but only in the reference list The diagram can be manipulated as described in section 2 4 2 The colored image can only be exported as a png file using the File Export image function in the applet menu i ivi SBGN Image View Activity Flow Figure 4 7 View results in pathways Each list id colored by a different color A total of 4 lists can be uploaded 4 1 2 Analyze gene list with expression values For each molecular function biological process cellular component PANTHER protein class or pathway term in PANTHER the genes associated with that term are evaluated according to the likelihood that their numerical values were drawn randomly from the overall distribution of values The Mann Whitney U Test Wilcoxon Rank Sum Test is used to determine the P value that say the chromatin packaging and remodeling genes have random values relative to overall list of values that were input This ap
41. d Whole Genome Pie Charts Section 2 4 1 from all 48 organisms e perform batch ID search see Section 3 3 e view statistics such as for each organism number of total genes number of genes in PANTHER families number of genes with GO classifications etc PANTHER EXE cones and orthotogs Trees and HMMs Pathways Ontologies Tools Workspace Keyword Search Batch ID Search PANTHER is giving a tutorial at the ICSB 2011 in Heidelberg on August 27 2011 Please click here for details Search GENES GENOMES AND ORTHOLOGS PANTHER now contains the complete sets of protein coding genes Genes and orthologs e for 48 organisms obtained from definitive sources Go For each gene PANTHER also reports orthologs and paralogs based on the inferred speciation and gene duplication events in the phylogenetic tree PANTHER also infers the least diverged orthologs LDOs which are the genes in two different organisms Whole genome function that have diverged the least since their common ancestor and are views most likely to retain the greatest functional similarities Quick links Gene expression tools cSNP tools Upload multiple gene IDs Keyword Search Batch ID Search Community Curation Search by keywords such as PANTHER Upload lists of IDs to to search by single or classification gene name gene symbol multiple accession numbers My Workspace protein accession HMM scoring Downloads P Whole Genome V
42. detailed description of the pathway Pathway Accession Components The number of pathway components within the pathway A pathway component is a group of homologous proteins across various organisms that participate in the same specific biochemical reactions within the pathway Clicking on the number will lead to a Pathway component list page section 2 2 4 Subfamilies The number of subfamilies that have at least one training sequence associated directly with the pathway Clicking on the number will lead to a Subfamily list page section 2 2 2 Associated Sequences The number of training sequences that are directly associated with the pathway by manual curation Clicking on the number will lead to the Gene list page section 2 2 1 References A list of literature references or websites used to generate the pathway diagram Author The author who generated the pathway diagram and curated training sequences to pathway associations Released On The date the pathway was generated and released CHAPTER 2 PANTHER AT A GLANCE 22 INSULIN IGF PATHWAY PROTEIN KINASE B SIGNALING CASCADE S3ML BGN els bes d eilDesigner Pathway Description Definition The insulin receptor IR insulin like growth factor receptor IGFR and insulin receptor related receptor IRR form a subgroup of receptor tyrosine kinases involved in regulating cellular growth and organismal metabolism The activated IR regulates cellular uptake and metabolism of fuel
43. e little icon right in front of the name On the top of the panel there are a few special utilities Search You can type the term in the Type the search term for live filtering box and only the ontology terms containing the search will be displayed Expand to selections Clicking this button will expand only the paths to the selected ontology terms Expand all Clicking this button will expand the ontology so that all child terms subcategories are shown Collapse all Clicking this button will collapse the ontology so that only the topmost general terms are shown Clear all selections Clicking this button will clear all selections of all types of on tologies Bookmark Clicking this button will cause the page to reload with a special address URL This address will have all your selection encoded If you save this address to your bookmark or favorites you can return to your current selection at any time This address is also suitable for cut and paste into your documents email messages or reference documentation e Results Type green arrow This panel is on the right side and it displays the number of genes PANTHER families and pathway molecule classes components that matches your selections You can choose one of the above result types and click the Get Results button The results will be displayed as a list page Section 2 2 e Selection Summary yellow arrow This panel is in the b
44. egicus Gallus gallus V Danio rerio V Drosophila melanogaster V Caenorhabditis elegans M Saccharomyces cerevisiae M Schizosaccharomyces pombe M Dictyostelium discoideum M Arabidopsis thallana M Escherichia coli CAN Figure 3 2 Advanced keyword search 13 2 Genes Close Customize genes criteria to search on M Training Sequence Entrez Gene ID fM Gene Name Gene symbol Alternate 1D PANTHER Family Subfamily PANTHER Molecular Function f PANTHER Blological Process PANTHER Cellular Component 4 PANTHER Protein Class M Speces 6 E Families Refine 6 kT Pathways Refine 25 T ontology terms Refine Figure 3 3 The search criteria can be refined by clicking the refine link for each subject The page is divided into 3 parts Fig 3 2 e The top part is the search box to enter the search term s e The middle part is the search results section It displays results from 4 subjects The number of results for each subject is on the left side red circle In the example in Figure CHAPTER 3 PANTHER BASICS 34 3 2 the search term of mdm2 yielded results of 13 genes 6 families 6 pathways and 25 ontology terms If you access this page from the keyword search link in Genes and orthologs main page section 2 1 1 it shows none in these brackets You can refine the search criteria by clicking the Refine link on the right side blue circle and make selections on the expanded
45. ens v Mus musculus v Rattus norvegicus v Gallus gallus v Danio rerio v Drosophila melanogaster v Caenorhabditis elegans v Saccharomyces cerevisiae v Schizosaccharomyces pombe v Dictyostelium discoideum v Arabidopsis thaliana v Escherichia coli Figure 3 6 Batch ID upload Please pay attention to the following e Supported IDs As mentioned earlier not all IDs are supported by the PANTHER We are working hard to improve this but at the moment only the following IDs are supported You can also click the supported IDs link to get the following list Please note that the system is not able to do partial match so you need to input the full IDs Ensembl Ensembl gene identifier Example ENSG00000126243 Ensembl PRO Ensembl protein identifier Example ENSP00000337383 Ensembl TRS Ensembl transcript identifier Example ENST00000391828 3 4 CHAPTER 3 PANTHER BASICS 38 Gene ID EntrezGene IDs examples include GeneID 10203 10203 for Entrez gene GeneID 10203 Gene symbol for example CALCA GE NCBI GI numbers Example 16033597 HGNC HUGO Gene Nomenclature ids Example HGNC 16673 PI International Protein Index ids Example IPI00740702 UniGene NCBI UniGene ids Examples Hs 654587 At 36040 UniProtK B UniProt accession Example 080536 UniProtK B ID UniProt ID Example AGAP3 HUMAN e File format The tool only accepts simple text txt in the following format If your
46. es that have protein kinase activity that involve in apoptosis pathway in the human genome 5 Human curators have associated ontology terms with genes PANTHER families subfam ilies and pathways To see these associations select the result type in the right panel and press the orange Get Results button blue arrow It will take you to the list pages Prowler Browse the PANTHER system using the Prowler and retrieve results for different data associated with the ontology and pathway terms such as individual genes or families and subfamilies of proteins About the PANTHER Ontologies PANTHER Browser RESULT pes ELT wo 5 Molecular Function ellular Component a E DNA directed RNA polymerase activity 32 Gene 24 PANTHER Family 14 Pathway molecule O E RNA directed DNA polymerase activity gt amp acyltransferase activity E B kinase activity O B amino acid kinase activity O E carbohydrate kinase activity E v Ed nucleotide kinase activity l fa E protein kinase activity Ed non membrane spanning protein tyrosi Ed transmembrane receptor protein kinase amp lt r Selection Summa All Gene results must belong to the following Molecular Function or its subcategories protein kinase activity and must also be involved in the following Pathway Apoptosis signaling pathway and must also belong to the following Species Homo sapiens
47. f Protein kinase receptor For GO terms the parent is not the parent category in the full Gene Ontology but the parent GO terms in the PANTHER GO Slim Clicking on the category name will lead to the Category Detail page Child The names of all child categories The names of all child categories A child category refers to a more specific category of the current one e g Serine threonine protein kinase receptor is a child category of Protein kinase receptor For GO terms the child is not the parent category in the full Gene Ontology but the child GO terms in the PANTHER GO Slim Clicking on the category name will lead to the Category Detail page Families and Subfamilies These 2 columns list the number of families and subfamilies that are classified by the category The yellow triangle in front of the column name indicates that the table is currently sorted by the column Genes and orthologs Trees and HMMs Pathways Ontologies Tools Workspace PANTHER ONTOLOGY TERMS LIST Convert List to Select Send list to Select B Display 30 E items per page Refine Search er E Category ID A Name Parent me Families Subfamilies O 31 GO 0006820 anion transport ion transport 13 849 QO 32 PCO00050 annexin calcium binding 20 816 protein O 33 GO 0009948 anterior posterior axis pattern specification 8 190 specification process QO 34 PCO0051 antibacterial response protein defense immun
48. f families and or subfamilies with the following related information Figure 2 7 e Family ID In this column it usually lists either family or subfamily IDs Click the ID will take you to the Family Detail page see section 2 3 2 e Family Name This column lists the names of families or subfamilies Click the name will take you to the Family Detail page see section 2 3 2 2 2 3 CHAPTER 2 PANTHER AT A GLANCE 12 e Genes This column lists the number of Genes in the family or subfamily These are also the sequences used to build the model Clicking on the number will take you to the Gene List page see section 2 2 1 e GO Molecular Function Biological Process Cellular Component These are Gene Ontology terms from PANTHER GO Slim describing the function of the gene prod uct You can click on each term to view more information in the Ontology Term Detail page see section 2 3 3 e PANTHER Protein Class This is a PANTHER Index terms describing protein classes You can click on each term to view more information in the Ontology Term Detail page see section 2 3 3 e Pathway This column lists the pathway and pathway component in which the families or subfamilies are involved Clicking on the terms will lead to the pathway diagram page section 2 4 2 If the pathway component is clicked the corresponding component will be highlighted in yellow when the diagram is open e Interpro If the PANTHER family or subfamily is
49. file contains a list of IDs separate the IDs by tab carriage return line return comma pipe or space If your file contains previously exported genelist see section 2 2 1 about how to export a gene list use the tab delimited file resulting from the export feature The system searches on the Gene ID column Microsoft Excel file format is not accepted by the tool To convert an Excel format xls or xlsx to a simple text format txt Open the file in Excel under File menu choose Save As and under the Format option choose Tab Delimited Text and then click the Save button 2 Select File Type As mentioned above you can select either ID list or Previously exported gene list 3 Results page You can choose what to display on the result page It can be either a Gene list page section 2 2 1 or protein list page 4 Select datasets This allows you to select what organism s you are searching Default is 12 model organisms PANTHER HMM Score The function can be accessed from the PANTHER home page or the interactive score of new sequences against PANTHER HMMs link on the Trees and HMMs subject home page section 2 1 2 This online functionality only allows you to submit one sequence at a time For multiple sequence scoring you need to download the PANTHER scoring tool from the PANTHER ftp site at http www pantherdb org downloads To score just enter the amino acid sequence into the box and click the Submit bu
50. g Specify color ranges for pathway diagram button on the result page Figure 4 10 First there are two color ranges to select Select equal CHAPTER 4 PANTHER TOOLS 52 distributions default and the ranges will be determined by sorting your gene values and then splitting the values into six equal distributions Select fold changes and standard gene expression fold change ranges will be used Some pathway components have multiple genes mapped to them You can she color ranges for these components that are determined by e median of the gene values default e average of the gene values e maximum gene value e minimum gene value File Edit Zoom Select Color amp Model Standard View Lit H Molecules Figure 4 12 Mann Whitney test results shown in pathway diagram in a head map CHAPTER 4 PANTHER TOOLS 53 gene expression color ranges Here you can specify how the pathway diagrams will be colored according to your gene expression values Select equal distributions and the ranges will be determined by sorting your gene values and then splitting the values into six equal distributions Select fold changes and standard gene expression fold change ranges will be used Select how to calculate the ranges Six equal distributions Fold changes 1 663E00 to 6 928E 01 6 928E 01 to 2 280E 01 2 280E 01 to 3 761E 02 3 761E 02 to 4 939E 02 4 939E 02 to 1 522E 01 Some pathway
51. generate the pathway dia gram CHAPTER 2 PANTHER AT A GLANCE 23 2 Sequence Association A list of all training sequences that are associated with the compo nent by manual curation The list includes the following columns PANTHER Subfamily PANTHER subfamily Ids It links to PANTHER subfamily information page Associated Sequences Training sequence ID It links to NCBI Entrez page Definition The gene definition for the training sequence Evidence Code See below Evidence A list papers with experimental results indicating the involvement of the se quence in the pathway It is usually in the form of PubMed IDs What is an Evidence Code Evidence Codes are used to denote the type of the evidence used for associating sequences to pathway components The PANTHER Evidence Code is modeled after the GO Evidence Codes http www geneontology org GO evidence shtml PANTHER Pathway gt p53 pathway Mdm2 PATHWAY COMPONENT DETAILS Name Mdm2 Definition MDM2 can inactivate several functions of the tumor suppressor p53 and can degrade p53 Category ID Synonyms Upstream Downstream Number Families Subfamilles Number of Associated Sequences GO Molecular Function GO Blological Process GO Cellular Component PANTHER protein class G01563 p53 Mouse double minute 2 Mdm2 3 11 References PubMed 14707284 PubMed 1614537 SEQUENCE ASSOCIATION Panther Subfamily Associated Sequence Def
52. iews Genomes and statistics View functions of all protein coding genes Genome statistics for Homo sapiens Mus musculus Rattus Site map norvegicus Gallus gallus Danio rerio Drosophila melanogaster Caenorhabditis elegans Saccharomyces cerevisiae Newsletter subscription Schizosaccharomyces pombe Dictyostelium discoideum Arabidopsis thaliana Escherichia coli Anopheles gambiae Aquifex aeolicus vf5 Ashbya gossypii Aspergillus nidulans Bacillus EX subtilis Bacteroides thetaiotaomicron Bos taurus Bradyrhizobium japonicum Caenorhabditis briggsae Canis familiaris Chlamydia trachomatis Chlamydomonas reinhardtii Chloroflexus aurantiacus Ciona intestinalis Deinococcus radiodurans Entamoeba histolytica Fugu rubripes Enter your Email Figure 2 1 Gene and orthologs home page 2 1 2 CHAPTER 2 PANTHER AT A GLANCE 7 Trees and HMMs PANTHER version 7 0 contains 6594 protein families each with a phylogenetic tree relating modern day genes in 48 organisms Phylogenetic trees also infer the evolutionary events that occurred such as speciation and gene duplication events Expert biologists have divided each family into subfamilies which are generally orthologous groups but may also contain recently duplicated paralogs Each family and subfamily is also represented as a hidden Markov model HMM which can be used to classify new sequences to an existing subfamily This page serves as a portal for users to
53. ificant p value indicates that the distribution for this category is non random and different than the overall distribution A cutoff of 0 05 is recommended as a starting point Results Analysis details Mapped IDs 25685 Unmapped IDs 3315 Specify color ranges for pathway diagrams Graph selected categories Export results Click on pathway name to see genes highlighted on pathway diagram Figure 4 10 The Mann Whitney test result page To have a visual representation of these distributions select the checkboxes of the categories of interest and click on the Graph selected categories button near the top of the page Figure 4 11 shows an example of the graph The x axis is your uploaded value The y axis is the 4 1 2 3 CHAPTER 4 PANTHER TOOLS 51 cumulative fraction In other words if you look at the data point x 1 4 y 75 this means that 75 of your uploaded values have a value of 1 4 or smaller 1 00 0 95 0 90 0 85 0 80 0 75 0 70 0 65 0 60 0 55 0 50 0 45 0 40 0 35 0 30 0 25 0 20 0 15 0 10 0 05 20 15 10 5 0 5 10 15 20 25 30 35 Uploaded value Tcell activation Overall Fraction Figure 4 11 Mann Whitney test results shown in graph view The Mann Whitney U Test Wilcoxon Rank Sum test Statistic To perform the rank sum test first the values of the genes that map to a given category are combined with the overall list of values that were input Then all the values are
54. in Clicking on the name will lead to the PANTHER family or subfamily detail page section 2 3 2 e GO Molecular Function Biological Process Cellular Component These are Gene Ontology terms from PANTHER GO Slim describing the function of the gene prod uct Clicking on the ontology term will lead to the term detail page section 2 3 3 CHAPTER 2 PANTHER AT A GLANCE 10 e PANTHER Protein Class This is a PANTHER Index terms describing protein classes Clicking on the term will lead to the term detail page section 2 3 3 e Pathway Pathway and pathway component with which at least one training sequence within the PANTHER best hit family or subfamily were associated directly by manual curation Clicking on the terms will lead to the pathway diagram page section 2 4 2 If the pathway component is clicked the corresponding component will be highlighted in yellow when the diagram is open e Species The organism of the gene hn te emt twee ea We NT PANTHER GENE LIST Customize Gene list Convert List to Select Send list to setect oO Display 30 items per page Refine Search Hits 1 26 of 26 page 1 Species Filter x x x E E Gene ID Gene Name Protein ID PANTHER GO Molecular GO Biological Process GO Cellular PANTHER Protein Pathway Species oer anj Gene Family Subfamily Function Component Class i Symbol 1 HUMAN ENSEMBL ENSGO0
55. inition Evidence Medline 7477327 Medline 7477327 Medline 8146175 Medline 8146175 Medline 8146175 PTHR10360 SF9 ENSBTAPO0000013766 null IGI Q00987 E3 ubiquitin protein ligase IGI Mdm2 P56950 E3 ubiquitin protein ligase IGI Mdm2 ENSBTAP00000013766 null IGI P23804 E3 ubiquitin protein ligase IGI Mdm2 Q561Z0 Mdm2 protein IGI PTHR10360 SF10 QSXINi Protein Mdm4 Iss PTHR10360 SF9 Q561Z0 Mdm2 protein IGI Q00987 E3 ubiquitin protein ligase IGI Medline 8146175 Medline 7477327 Medline 8146175 Mdm2 Figure 2 19 Pathway Molecule Class component detail page 2 4 2 4 1 CHAPTER 2 PANTHER AT A GLANCE 24 Graph and diagram pages Pie charts A pie chart can be generated from a gene list section 2 2 1 Whole genome pie charts for all 48 organisms in PANTHER have been pregenerated and can be accessed from the Genes and Orthologs subject home page section 2 1 1 Here is what you can do on this page Figure 2 20 e If it is a whole genome pie chart you can choose the ontology you want to display from the Select ontology drop down menu e From the Select species drop down menu you can choose one of the 48 organism you want to display e You can see the category names and statistics by mouse over the pie chart selection See detailed explanation of the statistics below e You can drill down to pie chart of child categories by clicking on a pie chart selection e You can retrieve a gene list by clic
56. integrated into Interpro the Interpro accession is listed here Clicking on the Interpro accession will lead you to the Interpro website PANTHER FAMILY LIST Convert List to Select 1 Send list to seec 1 Display 30 Items per page Refine Search GO Molecular Function E Biological Process r anj A FamiyiD Family Name Genes GO Cellular PANTHER Protein Pathway Interpro Component Class i PTHRi0000 PHOSPHOSERINE PHOSPHATASE 64 hydrolase activity acting on ester bonds cellular amino acid and derivative metabolic process phosphatase Serine glycine phosphatase activity biosynthesis gt Phosphoserine phosphatase C 2 PTHR10000 SFO PHOSPHOSERINE PHOSPHATASE 64 hydrolase activity acting on ester bonds cellular amino acid and derivative metabolic process phosphatase Serine glycine phosphatase activity biosynthesis gt Phosphoserine phosphatase CQ 3 PTHR10012 SERINE THREONINE PROTEIN 41 protein binding protein metabolic process phosphatase activator PHOSPHATASE 2A REGULATORY SUBUNIT phosphatase activator activit B phosphatase regulator activity 4 PTHRi0012 SF0 SERINE THREONINE PROTEIN 41 protein binding protein metabolic process phosphatase activator PHOSPHATASE 2A REGULATORY SUBUNIT phosphatase activator activit B phosphatase requlator activity 5 PTHR10026 SF24 SUBFAMILY NOT NAMED 4 DNA binding cell cycle nucleus transcription cofactor RNA bindi
57. ity 10 1068 protein O 35 GO 0003823 antigen binding binding 2 203 36 GO 0019882 antigen processing and immune system antigen processing and 6 320 presentation process presentation of peptide or polysaccharide antigen via MHC class II O 37 GO 0002504 antigen processing and antigen processing 4 260 presentation of peptide or and presentation polysaccharide antigen via MHC class II J 38 GO 0016209 antioxidant activity peroxidase activity 8 76 O 39 PCc00052 apolipoprotein transfer carrier 10 486 protein J 40 GO 0006915 apoptosis intracellular signaling negative requlation of 115 4082 cascade apoptosis induction of apoptosis Figure 2 10 Ontology term list page You can sort the list convert the list to a different ID type and save the list as described in Gene List section 2 2 Information detail Gene detail page This page provides detailed information about a gene The page contains 3 general sections 1 pages CHAPTER 2 PANTHER AT A GLANCE 16 1 General information about the gene Figure 2 11 Gene ID From source database either a model organism database e g FlyBase En sembl or Entrez Gene Protein ID A UniProt preferentially SwissProt ID whenever possible this is the rep resentative protein sequence for the gene Gene Name From source database together with the Gene ID Gene Symbol s Entrez gene symbol if available Organism The organism of the gene Alternative IDs Iden
58. k In the binomial test we assume that under the NULL hypothesis genes in the uploaded list are sampled from the same general population as genes from the reference set i e the probability p C of observing a gene from a particular category C in the uploaded list is the same as in the reference list We first estimate the probability p C from the reference set assuming that it is large and representative p C n C N where n C is the number of genes mapped to category C and N is the total number of genes in the reference set We then use the above estimate to find the p value the probability of observing k C genes or a more extreme number in the uploaded list of size K Under the NULL hypothesis the number of genes of mapped to C is distributed binomially with probability parameter p C and thus the p value would be K p vaue Y pofa e 5 where the sum runs from k C to K in the case of over representation i e when the number of observed genes k C is greater than expected p C K under the NULL hypothesis and 0 to k C in the case of under representation i e when k C is smaller than p C K When developing this analysis tool we tested using both the Chi Square and Binomial statistical tests We decided to use the Binomial since the Chi Square is not as accurate when the population sizes or the expect number is small View pathway diagram If you are analyzing pathways you can click the pathway name in the first
59. king on the chart legend link on the right side e The page also provides links to allow you to convert the pie chart to bar chart or to export the data as txt file so you can use your own program to create charts How to read the pie chart statistics When you mouse over the pie chart selection the category name and a series of numbers are displayed These numbers are the statistics for the category you selected Here is how the numbers are from 1 The first number is the number of genes that are classified to this category In our example in figure 2 20 it is 2067 2 The second number is the percent of genes classified to this category over the total number of genes Right above the pie chart it shows that the total number of genes is 17181 Remember this is the total number of human genes in the PANTHER library 3 The third number is the percent of genes classified to this category over total number of class hits in this case 13148 Class hit means independent ontology terms If a gene is classified to 2 ontology terms that are not parent or child to each other it counts as 2 class hits CHAPTER 2 PANTHER AT A GLANCE 25 PANTHER PIE CHART Bar Chart Export Features Mouse over pie chart section to see category name and statistics Click on a pie chart section to drill down to child categories Click on chart legend link to retrieve gene list for each category Select Ontology Protein Class H Select Species Homo
60. kspace Select the genes you want to save by clicking the check boxes The default is for all genes in the list You can select one of the followings from the pull down menu as the destination Figure 2 6 Workspace You need to register to save data to the workspace The registration is free When you make this selection a pop up window will ask you to name the list and add any comments The name and comments can be edited at any time in the future from the Workspace page Once the gene list is now at the site it can be returned to at any time Only the IDs are stored and they are mapped to the internal PAN THER gene ids so when you access a list in the future all information will be updated and current Exporting a list to a file The list will be exported as a tab delimited file You can now import the file into Excel or perform any post processing you wish View the list as text on the website e Use the pie chart view by clicking the colorful pie chart icon See Pie Charts section 2 4 1 for details PANTHER GENE LIST Customize Gene list Convert List to Select E Send list to J Works Display 30 F items per page Refine Search n space Hits 1 26 of 26 page 1 Text Figure 2 6 Export list 2 2 2 Family subfamily list page The family subfamily list page can be generated from prowler keyword search pathway dia gram and other list and detail pages This page provides a list o
61. lactose Figure 2 4 Gene list page Tip Manipulate your list Here is a description of what you can do to your list in any list pages in this section 2 2 e Sort the list You can always sort the list by clicking on any of the underlined column names A yellow triangle appears in front of the column name that you choose to sort The orientation of the triangle indicates the sort is ascending or descending e Customize columns You can click on the x button next to the column names to collapse the column e Converting a list to another list type Select the genes you want to convert by clicking the checkboxes The default is for all genes in the list Click on the pull down menu after Convert list to The current list type is shown in the box CHAPTER 2 PANTHER AT A GLANCE 11 Select the new list type from the pull down menu Figure 2 5 Each primary ID first column of the list is used to return the selected data type Note that the mapping between different types is not necessarily one to one e g a subfamily can map to more than one associated transcript PANTHER GENE LIST Customize Gene list Convert List to Select Send list to Select ch E J PANTHER Ontology Terms Display 30 PANTHER Families Hits 1 26 of 26 Pathways Pathway Components CHICK DANRE DROME RAT Figure 2 5 Options to convert a list from one type to another e Saving a list to your wor
62. ltiple sequence alignment a subPSEC score cannot be generated and the output will return the text string position does not align to the HMM indicating that the substitution occurs at a position that is inserted relative to the consensus HMM for the given HMM In most cases these positions are not modeled by the HMMs simply because they do not appear in most of the related sequences as a result substitutions at inserted positions are not generally likely to be deleterious EVOLUTIONARY ANALYSIS OF CODING SNPS Click on the link on the number of the multiple sequence alignment MSA position to vi the MSA where the substitution occurs 9 PANTHER HMM APOLIPOPROTEIN E PTHR18976 SF2 HMM E value score 2 2e 155 2 48955 0 37509 L46P 0 09251 0 0176 2 863 2 70376 0 42648 C130R 144 0 02918 0 1938 2 904 5 38922 0 916 R163C 177 0 39695 0 0029 2 904 5 10141 0 89104 R176C 19 0 34635 0 0035 2 904 The probability that a given variant will cause a deleterious effect on protein function is estimated by Pdeleterious such that a subPSEC score of 3 corresponds to a Pdeleterious Of 0 5 For more details please see Brunham et al 2005 Figure 4 15 cSNP analysis results page Appendix A Systems requirements e PC Users Recommended Windows 2000 or Windows XP Microsoft Internet Explorer 6 x Java version 1 4 2 is required e Macintosh Users A G4 Macinosh with MacOS 10 3 7 is required Safari versio
63. mily and subfamily is also Upload multiple gene IDs represented as a hidden Markov model HMM which can be used to classify new sequences to an existing subfamily Gene expression tools Community Curation My Workspace Search for PANTHER families trees and HMMs HMM scoring gt Browse families and subfamilies by classifications Downloads Interactively score a new sequence against PANTHER HMMs gt Download software to score many sequences against PANTHER HMMs Genome statistics Cita man Figure 2 2 Trees and HMMs home page CHAPTER 2 PANTHER AT A GLANCE 8 2 1 3 Pathways The current version of PANTHER Pathway consists of 165 pathways each with subfamilies and protein sequences mapped to individual pathway molecule class aka component All pathways are stored in both SBML and BioPAX format Pathway diagrams are interactive and include tools for visualizing gene expression data in the context of the diagrams This page provides a port for users to access information related to PANTHER pathways Figure 2 3 and provides the following links e Community Curation this link doesn t work since the site is under modification at the moment e Browse pathways this takes you to the prowler page section 3 2 e Search pathways this takes you to the advanced keyword search page section 3 1 2 e Pathway resources links to some useful pathway databases tools and resources e List of pathways
64. n nucleobase nucleoside nucleotide and nucleic acid metabolic process protein binding cell cycle kinase activator activity kinase regulator activity transcription factor activity transcription cofactor activity O 6 PTHR10029 ACYLPHOSPHATASE 61 hydrolase activity acting on ester bonds phosphate metabolic process phosphatase hosphatase activity C 7 PTHR10029 SF1 ACYLPHOSPHATASE 2 13 hydrolase activity acting on ester bonds phosphate metabolic process phosphatase phosphatase activity C 8 PTHR10029 SF2 ACYLPHOSPHATASE 1 10 hydrolase activity acting on ester bonds phosphate metabolic process phosphatase phosphatase activity 9 PTHR10029 SF3 ACYLPHOSPHATASE 30 hydrolase activity acting on ester bonds phosphate metabolic process phosphatase phosphatase activity Figure 2 7 Family subfamily list page You can sort the list convert the list to a different ID type and save the list as described in Gene List section 2 2 1 Pathway list page This page provides a list of pathways with the following information Figure 2 8 e Pathway Accession This column lists the unique accession of each pathway Clicking the accession will take you to the Pathway Diagram section 2 4 2 e Pathway Name This column lists the pathway names in the list Clicking the accession will take you to the Pathway Diagram section 2 4 2 2 2 4 CHAPTER 2 PANTHER AT A GLANCE 13 e Components This column lists the num
65. n 1 2 4 Java version 1 4 2 is required e For both Minimum of 128 MB RAM 256MB RAM recommended JavaScript Java applets and cookies must be enabled in your browser Java applet runtime parameters set to ms128m mx512m Xss16m Screen resolution of at least 800x600 dpi is strongly recommended 56 Bibliography 1 10 11 Huaiyu Mi Qing Dong Anushya Muruganujan Pascale Gaudet Suzanna Lewis and Paul D Thomas Panther version 7 improved phylogenetic trees orthologs and collabora tion with the gene ontology consortium Nucleic Acids Res 38 Database issue D204 10 1 2010 Huaiyu Mi and Paul Thomas Panther pathway an ontology based pathway database coupled with data analysis tools Methods Mol Biol 563 123 40 2009 Paul D Thomas Anish Kejariwal Nan Guo Huaiyu Mi Michael J Campbell Anushya Muruganujan and Betty Lazareva Ulitsky Applications for protein sequence function evo lution data mrna protein expression analysis and coding snp scoring tools Nucleic Acids Res 34 Web Server issue W645 50 7 2006 Paul D Thomas Giga a simple efficient algorithm for gene tree inference in the genomic age BMC Bioinformatics 11 312 2010 K Katoh K Kuma H Toh and T Miyata Mafft version 5 improvement in accuracy of multiple sequence alignment Nucleic Acids Res 33 2 511 8 2005 Raymond J Cho and Michael J Campbell Transcription genomes function Trends in Genetic
66. narily conserved part of the alignment is used to build HMM and it is called the match state which is designated with uppercase letters The other less conserved region is designated by lowercase letters and is called non match state If a sequence misses a position in the match state it is called a delete state and is designated by a dash If a sequence misses a position in the non match state it is called an insert state and is designated by a dot In the MSA menu of the MSA view you can make the following selections CHAPTER 2 PANTHER AT A GLANCE 31 e Percentage Identity for Entire Alignment This is the default selection and shows the alignment across the entire length of the all sequences e Percentage Identity for Match States This only shows the alignment for the match states The MSA uses uppercase letters to indicate match state positions lowercase letters to in dicate non match state positions dots to indicate insert state positions and dashes to indicate delete state positions PANTHER TREE VIEWER close window PANTHER Family Name MDM2 4 PTHR10360 Classification System Tree EEN Percentage Identity for Entire Alignment gt Grid MSA Percentage Identity for Match States sq VEAST v Subfamily conserved Mic SN Coloring for current sequences n HUMA GNVPDCRRTISAPHSSESMEQEKPCSLBEKRPRDGNIIRTGLVTCEH LKKAGASCPICKKEIQLVIKVPI SF10 RATIRGD GIVPDCRRTISA
67. nd All Subfamiliy nodes Protein Id sf name definition organism g Expand All Nodes MACMUIEN MOMA P53 BINDING PROTEIN MDM Macaca mul EI Reset Root to Main PANTRIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Pan troglody SF10 HU HUMANIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 Homo sapiens Et v Use Distances SF10 RATIRG RATIRGD MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 Rattus norv R SF10 MOUSEI MOUSEIMG MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 Mus musculus M Ladder Top SF10 B BOVINIENS MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 Bos taurus Ladder Bottom SF10 B BOVINIENS MDM4 P53 BINDING PROTEIN MDM4 Bos taurus Ladder Original CANFAIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Canis familia SF10 MONDOIENSEMB MONDOIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Monodelphis Color by Subfamily SF10 ORNANIENSEMBL ORNANIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Ornithorhyn SF10 CHICKIENTREZ 41 CHICKIENT MDM4 P53 BINDING PROTEIN MDM4 PREDICTED hypothetical pr Gallus gallus SF10 XENTRIENSEMBL E XENTRIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Xenopus tro SF10 DANREIZFIN ZDB GEI DANREIZFI MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 Danio rerio SF10 FUGRUIENSEMBL EN FUGRUIEN MDM4 P53 BINDING PROTEIN MDM4 Protein Mdm4 p53 binding pr Fugu rubripes EN ff SF9
68. now available in BioPAX level 3 July 13 2010 additional info Publications How to cite PANTHER PANTHER version 7 improved phylogenetic trees orthologs and collaboration with the Gene Ontology Consortium Mi et al Applications for protein sequence function evolution data mRNA protein expression nalysis and coding SNP scoring tools Thomas et al PANTHER a library of protein families and subfamilies indexed by function Thomas et al See complete publication list Below is an overview of the PANTHER home page the numbers correspond to those in Figure 1 1 1 Browse Links to the Prowler so that you can browse by functions pathways and species see Section 3 2 2 Genes and Orthologs Section 2 1 1 Links to the Gene Genome and Orthologs page CHAPTER 1 WELCOME TO PANTHER SYSTEM 3 10 11 12 13 14 15 16 1T 18 19 20 21 22 23 where you can perform simple Keyword Search Section 3 1 1 find Whole Genome Pie Charts Section 2 4 1 and upload batch IDs Section 3 3 Trees and HMMs Links to the page that provides links to functions related to trees and HMMs such as HMM scoring Section 3 4 download PANTHER HMM scoring program Section 3 5 etc Pathways Links to the page that contains pathway related knowledge Section 2 1 3 Ontologies Links to the page that provides information about the PANTHER GO Slim
69. nstructions for the creation of structured pathways that can be published in the PANTHER Pathway database Browse Pathways Browse a complete list of over 165 PANTHER pathways pathway components and view diagrams Search Pathways Search pathways and pathway components by keywords such as molecular function biological process pathway relationships and interactions or PANTHER families Pathway Resources See a list of useful resources including links to other pathway datasets and tools for editing and viewing biological models List of Pathways See a complete list of curated pathways Figure 2 3 PANTHER Pathway home page 2 2 2 2 1 CHAPTER 2 PANTHER AT A GLANCE 9 Ontologies This page provides a brief description of the ontolgoies used in PANTHER It also provides links to browse in prowler Section 3 2 and download the ontologies Tools PANTHER System is not only a database but also a data analysis platform that allows users to perform more advanced tasks and analyze research results using this integrated system This page provides links to the following tools e Gene expression data analysis tools see detail in Section 4 1 e Evolutionary analysis of coding SNPs see detail in Section 4 2 e PANTHER scoring see detail in Section 3 4 e Downloads see Section 3 5 Workspace The Workspace is a unique feature in PANTHER that allows users to store the gene lists that they generate for future analysi
70. ntal data analysis 3 As mentioned in the toool home page section 2 1 5 there are three categories of tools gene expression analysis tools coding SNP analysis tools PANTHER scoring tool PANTHER scoring tool has already been mentioned in section 3 4 This chapter will describe the rest of the tools Gene expression data analysis tools Compare Gene List Use the binomial statistics tool to compare classifications of multiple clusters of lists to a reference list to statistically determine over or under representation of PANTHER classification categories Each list is compared to the reference list using the binomial distribution test Cho and Campbell TIGs 2000 6 for each molecular function biological process cellular component PANTHER protein class or pathway term in PANTHER Here is how it works e PANTHER select specific data source s for the genome of each organism It uses UniProt idmapping mechanism to map to a number of other IDs see supported IDs e The tool maps the IDs from your uploaded file to the IDs in the PANTHER database e Since all IDs in the PANTHER database map to the IDs we use to build PANTHER protein families thus your IDs are mapped to our PANTHER families and subfamilies and carry over all the ontology and pathway classifications e The tool will then count the number of your IDs in each of the category compare with the reference list and use binomial distribution to calculate the p value Bel
71. o they can track the identifier on the PANTHER website e The second column should be corresponding PANTHER ID ex PTHR10078 or PCTHR10078 SF 1 and is used to look up the molecular function biological process and pathway associations e The third column must be the corresponding numerical value Steps 1 Upload gene expression data file 1 Upload file File must be tab delimited File type Gene Transcript Protein and Alternate ID yP Upload file Users mi Documents PA Browse J supported IDs Figure 4 8 Upload a gene list with expression values After you select the file click the Upload file button On the next page you can select the ontology or pathway to analyze Figure 4 9 Steps 1 Upload gene expression data file 1 Upload file File must be tab delimited gt 2 Select search options Uploaded file genexpSample Warnings e 2 Search options wp PANTHER Ontology duplicates the Pathways first Id value pair O GO Biological Process In the file will be O GO Molecular Function used O GO Cellular Component O PANTHER Protein Class Figure 4 9 Select a PANTHER ontology or pathway to analyze 4 1 2 2 Results On the results page there is a table with four essential columns of data CHAPTER 4 PANTHER TOOLS 50 Pathways AP value E T cell activation 162 3 84E 04 C Glutamine glutamate conversion 6 5 90E 04 C Transcription requlation by bZIP tran
72. ods Mol Biol 563 123 40 PANTHER software and tools 3 Thomas PD Kejariwal A Guo N Mi H Campbell MJ Muruganujan A Lazareva Ulitsky B 2006 Applications for protein sequence function evolution data mRNA protein ex pression analysis and coding SNP scoring tools Nucleic Acids Res 34 Web Server issue W645 50 PANTHER tree building algorithm GIGA 4 Thomas PD 2010 GIGA a simple efficient algorithm for gene tree inference in the genomic age BMC Bioinformatics 11 312 1 3 PANTHER help This manual is not the only help you can get to learn how to use the PANTHER System You can get additional help and support through the following e On most of our webpages there are links to help tips that provide detailed information about the page and how to use it Simply click the 9 icon to get to the Tips page e You can always contact us through our feedback email at feedback pantherdb org 1 4 CHAPTER 1 WELCOME TO PANTHER SYSTEM Overview OK here we go The PANTHER home page Figure 1 1 provides an overview of the PAN THER functionality The top menu bar item 1 7 in Figure 1 1 contains links to individual subject main pages with links within that subject The left panel item 11 20 in Figure 1 1 con tains links directly to the popular tools for quick access The right panel contains PAN THER News and Publications item 23 and 24 respectively in Figure 1 1 The Keyword Search and Sequence Search HMM scoring
73. oteins as described in Thomas et al 2003 and Thomas amp Kejariwal 2004 Enter a protein sequence X2 Enter substitution s e g A265v 2 L46P R130C R163C C176R Figure 4 14 Enter sequence and SNP substitutions Note that the sequence numbering must be correct relative to the sequence pasted in the box and not necessarily the numbering found in the literature For example APOEA is listed as containing C112R in the literature but this requires removing the signal peptide from the SWISS PROT entry for APE HUMAN or entering the substitution as C130R Results of cSNP analysis tool Protein score The input protein sequence is scored against the HMMs in the PANTHER library The align ment to the HMM with the most significant score is used for the analysis Proteins that scored greater than 1e 23 are excluded from the analysis as the alignments are less reliable SubPSEC score and Pdeleterious The subPSEC substitution position specific evolutionary conservation score estimates the like lihood of a functional effect from a single amino acid substitution It is the negative logarithm of the probability ratio of the wild type and mutant amino acids at a particular position PAN THER subPSEC scores are continuous values from 0 neutral to about 10 most likely to be deleterious 3 is the previously identified cutoff point for functional significance A cutoff of 3 corresponds to a 5096 probability that a
74. ottom of the Prowler Since you are allowed to make selections from multiple ontologies a summary of the query results from your selections is displayed in this panel Browse the PANTHER ontologies Figure 3 5 shows an example of making selections using the PANTHER Prowler 1 Select an ontology on the Ontology type panel 2 Select the ontology you are interested in left panel Typing in the text field will filter the list of possible selections Click on a red arrow to expand a category to view subcategories and select categories by clicking on the checkbox next to the name Brief details about each category can be seen by mousing over a name or full details by clicking on the icon to the left of a name For pathways full details include a detailed diagram of the molecular interactions and reactions in the pathway CHAPTER 3 PANTHER BASICS 36 3 More than one ontology can be selected When multiple selections are made it means that results will meet the criteria from all the selections In this example selections are made in both Molecular Function and Pathway and the selected ontology can be seen as marked in the Ontology panel red oval in Figure 3 5 4 Review your selections in the Selection Summary panel green arrow For example in this example one selects Human from the species ontology as well as Protein kinase activity from the molecular function ontology and Apoptosis signaling pathway in pathway to retrieve all gen
75. ow is a step by step description of how to use the tool Select lists to analyze When you enter the tool the first step is to click Select file s button to upload the list s you would like to analyze Figure 4 1 You may upload up to four lists There are two ways you can upload the list s 1 Browse and upload from your computer Figure 4 2 This is similar to Batch ID Search described earlier section 3 3 Please pay attention to the following e Supported IDs As mentioned earlier not all IDs are supported by the PANTHER We are working hard to improve this but at the moment only the following IDs are supported You can also click the supported IDs link to get the following list 42 CHAPTER 4 PANTHER TOOLS 43 Please note that the system is not able to do partial match so you need to input the full IDs Ensembl Ensembl gene identifier Example ENSG00000126243 Ensembl PRO Ensembl protein identifier Example ENSP00000337383 Ensembl TRS Ensembl transcript identifier Example ENST00000391828 Gene ID EntrezGene IDs examples include GeneID 10203 10203 for Entrez gene GeneID 10203 Gene symbol for example CALCA GI NCBI GI numbers Example 16033597 HGNC HUGO Gene Nomenclature ids Example HGNC 16673 IPI International Protein Index ids Example IPI00740702 UniGene NCBI UniGene ids Examples Hs 654587 At 36040 UniProtK B UniProt accession Example 08053
76. owing PANTHER data and tools e PANTHER HMM library The current version is 7 0 that contains the PANTHER HMMs for 6594 protein family HMMs divided into 62 972 functionally distinct protein subfamily HMMs The PANTHER HMMs are in the HMMER format There is also a version of the library available that can be used for the Coding SNP Analysis tool e PANTHER HMM scoring tool Download the PANTHER scoring tool to score protein CHAPTER 3 PANTHER BASICS 40 sequences against the entire PANTHER HMM library and analyze your sequences The HMM library should be downloaded separately e PANTHER cSNP scoring tool Download the PANTHER Coding SNP Analysis tool to estimate the likelihood that a particular nonsynonymous amino acid changing coding SNP will cause a functional impact to the protein A special version of HMM library for this tool should be downloaded separate e GIGA GIGA is an efficient tree building program that allows phylogenetic reconstruction of very large gene families and determination of orthologs on a large scale It is used to generate trees in the PANTHER Classification System e PANTHER Pathway Contains 165 regulatory and metabolic pathways each with sub families and protein sequences mapped to individual pathway components All pathway diagrams are abailable in both SBML and BioPAX level 3 format SBML files BioPAX files and Protein Sequence Association data are available for download e PANTHER Sequence Classifi
77. panel Figure 3 3 e The bottom part allows you to select genome s from the 12 Model Organisms that you want to search The default is all 3 2 Prowler The PANTHER Ontology Browser or PANTHER Prowler is a highly interactive Java applet used to browse the PANTHER ontologies make selection s and retrieve results for different data associated with the ontology terms such as individual genes or families and subfamilies of proteins The Ontology Browser is composed of 4 panels Figure 3 4 2 Prowler Browse the PANTHER system using the Prowler and retrieve results for different data associated with the ontology and pathway terms such as individual genes or families and subfamilies of proteins About the PANTHER Ontologies Result Types foe E ocn E apoptosis E cell adhesion 0 PANTHER Family Ed cell communication j 0 Pathway molecule E cell cycle E3 cellular component organization E3 cellular process Ed developmental process Ed generation of precursor metabolites and energy E3 growth Ed homeostatic process Ed immune system process PANTHER Browser acc o Biological Proc Molecular Function Cellular Component Protein Class Pathway Species E E E E E E EE EE EE HOOOOOOOoOoOoO Your Selection Summary No selections have been made Quick Prowler tutorial for the first time user The Prowler is a Java applet used to browse the PANTHE
78. proach has been used by our group Clark et al 2003 7 and is similar to a method from Eric Lander s group Mootha et al 2003 8 to find weakly coordinated shifts that elude methods based on defining strict cutoffs in the data e g only focusing on genes whose expression has changed by over 1 5 or 2 fold For the rank sum test it is important to provide values for as many genes as possible subject to noise level and reliability so that randomness can be properly assessed across the experiment In most cases you should upload the entire list of the expression experiment CHAPTER 4 PANTHER TOOLS 49 4 1 2 1 Upload gene expression file When you enter the tool you can upload your gene expression file from your computer to the system using the interface as shown in Figure 4 8 The uploaded file must be a tab delimited text file and must contain an identifier to be analyzed and the corresponding numerical value You must select the file type to specify what type of identifier ID you are uploading If your file uses one of the supported IDs you select Gene transcript protein and alter native ID option and the file must contain tow columns The first column is the ID and the second is the numerical value If you use the PAN THER Generic Mapping File the file should contain 3 columns e The first column can contain any arbitrary ID but most be unique since this allows the user to uniquely specify each record in the dataset s
79. re those involved in the reaction that is controlled or modulated by the current component Clicking it lead to the Pathway molecule class detail page section 2 3 5 e Pathway The pathway that the component belongs to Clicking it lead to the Pathway molecule diagram page section 2 4 2 e Associated Sequences This column lists the numbers of genes that are associated to this pathway component Clicking the number will take you to the Gene list page section 223 e Subfamilies This column provides the number of subfamilies of all genes that are associ ated to the pathway component Clicking the number will take you to the Subfamily list page section 2 2 2 e GO classifications and PANTHER protein class These 4 columns lists all the categories classified to the genes that are associated to the pathway component Clicking on an ontology terms will lead to the Ontology Term Details page section 2 3 3 PANTHER PATHWAY COMPONENT LIST Convert List to Select E Sendlistto seec i Display 30 B items per page Refine Search Hits 1 20 of 20 page 1 cen an A Component Type Upstream Downstream Pathway Associated Subfamilies GO Ico GO Cellular PANTHER Component Name Sequence Molecular Biological Component Protein Accession Function Process Class Oo G04674 Mdm2 Gene p53 Mdm2 Insulin IGF pathway pro
80. rms the parent is not the parent category in the full Gene Ontology but the parent GO category in the PANTHER GO Slim Clicking the name will take you to the detail page of the ontology term CHAPTER 2 PANTHER AT A GLANCE 21 Child The list of names of all child categories For GO terms the child is not the child category in the full Gene Ontology but the child GO terms in the PANTHER GO Slim Clicking the name will lead you to the PANTHER detail page of the child category Number of subfamilies Families The number of subfamilies and families that are classified by the category Clicking on the number will lead to the Family Subfamily List page section 2 3 2 PANTHER CLASS INFORMATION Name Definition 6 ISBN 0198506732 Oxford Dictionary of Biochemistry and Molecular Biology Class ID GO 0016874 Parent Lineages molecular function catalytic activity ligase activity Child DNA ligase activity aminoacyl tRNA ligase activity ubiquitin protein ligase activity Number Subfamilles Families 2638 Figure 2 17 Ontology term detail page 2 3 4 Pathway description page This page provides details of the pathway To reach this page just click the pathway name link anywhere on the PANTHER website and then click the Pathway Description tab on the page You can toggle between this page and Pathway Diagram page This page provides following information about a pathway Figure 2 18 Definition A
81. s 16 9 409 415 2000 A G Clark S Glanowski R Nielsen P D Thomas A Kejariwal M A Todd D M Tanenbaum D Civello F Lu B Murphy S Ferriera G Wang X Zheng T J White J J Sninsky M D Adams and M Cargill Inferring nonneutral evolution from human chimp mouse orthologous gene trios Science 302 5652 1960 3 12 2003 Vamsi K Mootha Cecilia M Lindgren Karl Fredrik F Eriksson Aravind Subramanian Smita Sihag Joseph Lehar Pere Puigserver Emma Carlsson Martin Ridderstr le Esa Laurila Nicholas Houstis Mark J Daly Nick Patterson Jill P Mesirov Todd R Golub Pablo Tamayo Bruce Spiegelman Eric S Lander Joel N Hirschhorn David Altshuler and Leif C Groop Pgc lalpha responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes Nat Genet 34 3 267 13 7 2003 L R Brunham R R Singaraja T D Pape A Kejariwal P D Thomas and M R Hay den Accurate prediction of the functional significance of single nucleotide polymorphisms and mutations in the abcal gene PLoS Genet 1 6 e83 12 2005 Paul D Thomas Michael J Campbell Anish Kejariwal Huaiyu Mi Brian Karlak Robin Daverman Karen Diemer Anushya Muruganujan and Apurva Narechania Panther a library of protein families and subfamilies indexed by function Genome Res 13 9 2129 41 9 2003 Paul D Thomas and Anish Kejariwal Coding single nucleotide polymorphisms associated wi
82. s Although users do not have to register to use the PANTHER system registration is required in order to user the workspace Registration is free This webpage allows registered users to login or retrieve password and for non registered users to register A more detailed description of using Workspace can be found in Section 3 6 List pages Gene list page Thge gene list page can be generated from the prowler keyword search pathway diagram and other list and detail pages The gene list page contains the following information Figure 2 4 e Gene ID This is the identifier for genes in the PANTHER library The format is as follows organism gene database source gene id protein database source protein id For example HUMAN ENSEMBL ENSGO00000111262 UniProtK B Q09470 is a human se quence the gene sequence is from ENSEMBL with id ENSG00000111262 and the protein sequence is from UniProt with id Q09470 Clicking on the ID will lead to the Gene Detail page section 2 3 1 e Gene Name Gene Symbol The Entrez gene definition and gene symbol Clicking on the gene symbol will lead to the Gene Detail page section 2 3 1 Please note that some of the genes do not have the information in this column e Protein ID The public protein identifier used in the GeneID above Clicking on the ID will lead to the Gene Detail page section 2 3 1 e PANTHER Family Subfamily The family or subfamily name of the PANTHER model where the sequence is
83. s while the activated IGFR promotes cell growth survival and differentiation Knockout mice studies have revealed the IGF I receptor mediates IGF I and IGF II action on prenatal growth as well as IGF I action on postnatal growth The IR mediates prenatal growth in response to IGF II and postnatal growth in response to insulin All of these receptors share extensive structural homology and utilize the same downstream signalling cascades one of which is illustrated here Thus how specificity is achieved in physiological outputs remains unclear although it is thought to occur in part by differences in the locations and levels of receptor expression Pathway Accession P00033 Components 18 Families Subfamilies 5 Associated Sequences 275 References PubMed 11737239 PubMed 11897402 PubMed 12175645 PubMed 12360255 PubMed 11739335 PubMed 12169433 Author Huaiyu Mi Arnie Levine Released On Figure 2 18 Pathway Description page 2 3 5 Pathway molecular class component detail page A pathway component represents a group of homologous proteins across various organisms that participate in the same specific biochemical reactions within the pathway A pathway component is always associated with genes within the PANTHER libraries thus it is linked directly or indirectly to the PANTHER family subfamily statistical models This page provides details of a pathway component in two separate parts Figure 2 19 1 Pathway component
84. s a list of molecules shown in the diagram The types of the molecules are in parentheses e g PROTEIN SIMPLE MOLECULE GENE etc e The right panel is the diagram All objects on the diagram are interactive so you can customize the diagram according to your preferences The modified diagram can be saved as png file as described in section 2 4 2 2 There are three views for the diagram e Interactive Standard Activity Flow When this tab is selected the applet allows you to view two views that are not SBGN compliant CHAPTER 2 PANTHER AT A GLANCE 26 Standard View This view faithfully reflects the original diagram generated by the CellDesigner software It captures detailed molecular events or biochemical reactions within the pathway It is a little different from the conventional way of illustrating pathways For example if a protein A activates protein B it will be illustrated as protein A catalyzing a transition reaction of protein B from a non active to an active form Activity Flow This view uses simpler or more conventional relationship notations to illustrate information flowchart of the pathway It captures mainly 4 relationships between 2 proteins activation inhibition neutral and unknown It is derived based on the Standard View In cases where a reaction can not be simply illustrated as one of the above four it will keep it in the original Standard view This view is not compliant with SBGN Activity Flow
85. sapiens B PANTHER Protein Class Click to get gene list for a category Total Genes 17181 Total protein class hits 13148 Wi calcium binding protein PCO0060 chaperone PC00072 toskeletal protein PCO0085 efense immunity protein PC00090 nzyme modulator PCO0095 extracellular matrix protein PC00102 hydrolase PC00121 isomerase PC00135 igase PC00142 ase PC00144 membrane traffic protein PC00150 nucleic acid binding PC00171 oxidoreductase PC00176 hosphatase PC00181 rotease PC00190 receptor PC00197 signaling molecule PC00207 storage protein PC00210 Chart tooltips are read as Category name Accession genes Percent of structural protein PCO0211 gene hit against total genes Percent of gene hit against total Protein surfactant PC00212 transcription factor PC00218 transfer carrier protein PC00219 transferase PC00220 Wi transmembrane receptor requlatory adaptor protein PC00226 Wi transporter PC00227 E viral protein PC00237 ca 2 i gt Q amp p S5 2 3B e o 58 E o O ei S8 N o Jo jov re transcription factor PCO0218 2067 12 0 15 7 i Figure 2 20 The pid chart of the entire human genome in PANTHER Protein Class 2 4 2 Pathway diagram 2 4 2 1 PNATHER Pathway Applet Basics The pathway diagram on this page is interactive and is displayed in an applet window The window is divided into 2 panels Figure 2 21 e The left panel show
86. score is deleterious From this the probability that a given variant will cause a deleterious effect on protein function is estimated by Pdeleterious such that a subPSEC score of 3 corresponds to a Pdeleterious of 0 5 For more details and to see the corresponding equations please see our most recent pub lication Brunham et al 2005 9 Additional information is in Thomas et al 2003 10 and Thomas and Kejariwal 2004 11 Multiple sequence alignment Click on the link on the number of the multiple sequence alignment MSA position to view the column in the MSA where the substitution occurs The column is highlighted in red and the subfamilies rows in the MSA that were used to gather the statistics for amino acid probabilities 4 2 2 4 CHAPTER 4 PANTHER TOOLS 55 are highlighted in blue In the MSA view click on a blue diamond a subfamily node in the tree to hide or show sequences in the subfamily Number of independent counts NIC NIC number of independent counts is an estimate of the number of independent observations used to calculate the amino acid probabilities The probabilities are calculated from a combi nation of prior knowledge e g that isoleucine often substitutes for valine and observations so the larger NIC the more the probabilities rely on the amino acids observed in the multiple sequence alignment position does not align to the HMM If the substitution occurs at a position that does not appear in the mu
87. scribing protein classes PANTHER TREE VIEWER close window PANTHER Family Name MDM2 4 PTHR10360 Classification System Tree MSA 4 Protein Id sf name definition organism gene id gene symbol Ortho Molecular Biological Cellular co Protein Cla MACMUIEN MDM4 P53 BIND Protein Mdm4 p53 b Macaca mul ENSEMBLE GO 001687 GO 000691 PCO00142 lig a MACMUIEN MDM4 P53 BIND Macaca mul ENSEMBL E GO 001687 GO 000691 PC00142 lig PANTRIEN MDMA4 P53 BIND Protein Mdm4 p53 b Pan troglody ENSEMBLE GO 001687 GO 000691 PCO00142 lig HUMANIEN MDM4 P53 BIND Protein Mdm4 Homo sapiens ENSEMBL E MDM4 GO 001687 GO 000691 PC00142 lig pecs paene peores pones peer Rok GO 0016874 ligase activity GO 0004842 ubiquitin protein ligase activit MOUSEIMG MDM4 P53 BIND Protein Mdm4 Mus musculus MGI MGIz1 Mdm4 G UTU TO VU BOVINIENS MDM4 P53 BIND Protein Mdm4 Bos taurus ENSEMBLE MDM4 GO 001687 GO 000691 PC00142 ig BOVINIENS MDM4 P53 BIND Bos taurus ENSEMBLE GO 001687 GO 000691 PC00142 lig CANFAIEN MDM4 P53 BIND Protein Mdm4 p53 b Canis familia ENSEMBLE GO 001687 GO 000691 PC00142 lig MONDOIEN MDM4 P53 BIND Protein Mdm4 p53 b Monodelphis ENSEMBLE GO 001687 GO 000691 PC00142 ig ORNANIEN MDM4 P53 BIND Protein Mdm4 p53 b Ornithorhyn ENSEMBL E GO 001687 GO
88. scription Factor S6 8 23E 04 C p53 pathway 113 8 81E 04 C General transcription regulation 41 1 96E 03 C Inflammation mediated by chemokine and cytokine signaling pathway 315 3 28E 03 1 The first column contains the name of the PANTHER classification category If you are doing this analysis in terms of pathways you can click on the pathway name to view the pathway diagram see below The genes in the pathway diagram are colored according to the gene expression value and the rules for this can be specified by clicking on the Specify color ranges button The second column contains the number of genes that map to this particular PANTHER classification category The third column has either a or A plus sign indicates that for this category the distribution of values for your uploaded list is shifted towards greater values than the overall distribution of all genes that were uploaded A negative sign indicates that the uploaded list is shifted towards smaller values than the overall list The fourth column contains the p value as calculated from the Mann Whitney U Test Wilcoxon Rank Sum test A large p value indicates that the genes for this category have a distribution that is similar to randomly choosing genes from the overall distribution In other words the values of the uploaded genes for this category have a similar distribution to the overall list of values that were input A small sign
89. sis Please login to select lists from your workspace Figure 4 4 Upload is completed Steps 1 Select Lists to Compare to a Reference List 1 Select list s to For example each selected list may be a cluster of co expressed genes under a particular analyze set of conditions gt 2 Select reference list Select list s selected genexpSample 1 2 Select Reference List For example the reference list may be the set of all genes in the experiment or the set of all genes in the genome being analyzed Select reference list default Homo sapiens genes Search options PANTHER Ontology Pathways O Biological Process O Molecular Function O GO Cellular Component O PANTHER Protein Class Figure 4 5 Modify reference list if needed and then start the analysis 4 1 1 3 Analysis results On the results page there is a table with six essential columns of data 1 The first column contains the name of the PANTHER classification category If you are doing this analysis in terms of pathways you can click on the pathway name to view the corresponding pathway diagram see section 4 1 1 4 below 2 The second column contains the number of genes in the reference list that map to this particular PANTHER classification category CHAPTER 4 PANTHER TOOLS 46 3 The third column contains the number of genes in your uploaded list that map to this PANTHER classification category The fourth column contains the expected value whi
90. species e The LDO can be loosely interpreted as the most nearly equivalent gene in another organism If there were no gene duplication events following the speciation event that separated two species there is only a single ortholog in each genome and they are therefore marked as LDO If there was a gene duplication event following speciation there may be more than one ortholog and the least diverged ortholog pair is marked LDO while other ortholog pairs are marked O CHAPTER 2 PANTHER AT A GLANCE 18 ORTHOLOGS ID Organism Type HUMAN ENSEMBL ENSG00000198625 UniProtKB 015151 Chordata Osteichthyes P PANTR ENSEMBL ENSPTRG00000005203 ENSEMBL ENSPTRPOO000008832 Pan troglodytes LDO MACMU ENSEMBL ENSMMUGO0000014193 ENSEMBL ENSMMUP00000018653 Macaca mulatta LDO MOUSE MGI MGI 96952 UniProtKB P23804 Mus musculus LDO RAT RGD 1305332 NCBI XP 235169 Rattus norvegicus LDO BOVIN ENSEMBL ENSBTAGO0000026775 ENSEMBL ENSBTAPO0000013766 Bos taurus o BOVIN ENSEMBL ENSBTAG00000010422 UniProtKB ASPJWS Bos taurus LDO CANFA ENSEMBL ENSCAFG00000000418 UniProtKB P56950 Canis familiaris LDO MONDO ENSEMBL ENSMODGO00000007219 ENSEMBL ENSMODPO00000008952 Monodelphis domestica LDO ORNAN JENSEMBL ENSOANGO00000005942 ENSEMBL ENSOANPO00000009460 Ornithorhynchus anatinus LDO CHICK ENTREZ 395609 NCBI XP 416084 Gallus gallus LDO XENTR ENSEMBL ENSXETGO0000001434 UniProtKB Q6P3Q9 Xenopus tropicalis LDO FUGRU ENSEMBL
91. t PANTHER Ontology Terms A list of PANTHER Molecular Function and Biological Process ontology terms that are classified to PANTHER family or subfamily models with at least one training sequence associated with the selected pathway component PANTHER Families A list of PANTHER families or subfamilies with at least one training sequence associated with the selected pathway component Pathway Pathway Components 2 4 3 2 4 3 1 CHAPTER 2 PANTHER AT A GLANCE 28 TreeViewer PANTHER trees allow you to explore the relationships between sequences in a particular family as well as view some of the key information that was used to annotate the families and subfam ilies The current PANTHER release version 7 0 contain publicly available protein sequences from UniProt Refseq ENSEMBL and Model Organism Databases The Tree Viewer has two panels that are mapped to each other Figure 2 22 The left panel displays the relationship between the different sequences and is labeled as Tree The right panel has two views The first is the attribute table which is labeled as Grid and contains information for the each corresponding sequence in the tree panel The second view is the multiple sequence alignment view which is labeled as MSA Figure 2 24 You can click the the button to toggle between the two views Trees In the tree panel on the left of the Tree Viewer you can explore that phylogenetic relationships among different genes
92. t protein and substitution data 4 2 2 Results of cSNP analysis tool A Systems requirements 22 24 24 25 28 32 32 32 33 34 36 38 39 40 42 42 42 48 53 53 54 56 1 1 1 2 Chapter 1 Welcome to PANTHER System About this document This manual provides a step by step instruction of how to use all the pages functions and tools in the PANTHER Classification System website The manual starts with a detailed description of different types of data pages Chapter 2 followed by Chapter 3 which describes the basic functionalities of the system Chapter 4 provides detailed description of our data analysis tools This manual does not serve as a scientific reference of the PANTHER System It assumes that you are already familiar with the scientific background of the system If not we highly encourage you to read the papers listed in the next section to get familiar with our system How to cite PANTHER Here is a list of PANTHER publications based on the subject that you can cite as references when you publish your results e General PANTHER usage 1 Mi H Dong Q Muruganujan A Gaudet P Lewis S Thomas PD 2010 PANTHER ver sion 7 improved phylogenetic trees orthologs and collaboration with the Gene Ontology Consortium Nucleic Acids Res 38 Database issue D204 10 PANTHER pathway 2 Mi H Thomas P 2009 PANTHER pathway an ontology based pathway database cou pled with data analysis tools Meth
93. tation for all sequence below the collapsed node is shown in the appropriate columns Under the Tree menu you can do the followings Figure 2 22 e Collapse lowest level of Subfamilies A PANTHER subfamily node can have other sub family node s as descendants This option collapses all subfamily nodes that do not contain another subfamily node All subfamily nodes that nest another subfamily node remain expanded e Expand all Subfamily nodes All subfamily nodes are expanded All non subfamily nodes remain unchanged CHAPTER 2 PANTHER AT A GLANCE 29 e Expand all Nodes All nodes including non subfamily nodes can be expanded e Use distances If this option is selected a check mark shows in front of the option the tree branches are shown based on the distances calculated in tree building algorithm GIGA 4 If this option is unchecked the tree branch will be shown with fixed length e Ladder Top Re arrange the tree so that the node with more descendants are always above the nodes with less e Ladder Bottom Opposite to the above Re arrange the tree so that the node with more descendants are always below the nodes with less e Latter Original Set the order of the tree to what was loaded from the server It is different from either Ladder Top or Ladder Bottom e Colore Subfamilies PANTHER TREE VIEWER close window Family Name MDM2 4 PTHR10360 Collapse lowest level Subfamilies MSA Expa
94. tein kinase B signaling cascade 11 3 gt O 2 04675 PTEN Gene p53 Phosphatase and tensin homolog Insulin IGF pathway protein kinase B signaling cascade 19 Z Q 3 G04676 pii0alpha Gene p53 P110alpha Insulin IGF pathway protein kinase B signaling cascade 10 i Q 4 P00322 Cytochrome Protein Apoptosis protease activating factor 1 Apoptosis signaling pathway 54 2 gs P00894 AKT Protein 3 phosphoinositide dependent protein kinase Forkhead transcription factor Insulin IGF pathway protein kinase B signaling cascade 13 by Tuberin O 6 00895 INSRIGF R Protein Insulin receptor substrate famil Insulin IGF pathway protein kinase B signaling cascade 28 Zz IRR OQ 7 P00896 Insulin IGF Protein Insulin receptor substrate famil Insulin IGF pathway protein kinase B signaling cascade 32 4 O 8 00897 SHIP2 Protein Insulin IGF pathway protein kinase B signaling cascade 5 1 oe PO0898 FKHR Protein Protein kinase B Insulin IGF pathway protein kinase B signaling cascade 7 1 10 P00899 IRS 1 4 Protein Insulin receptor Insulin like growth factor receptor insulin related receptor Insulin IGF pathway protein kinase B signaling cascade 15 4 Insulin Insulin like growth factor 11 P00900 PI3 K Protein Insulin IGF pathway protein kinase B signaling cascade 95 18 12 P00902 GSK 3 Protein Protein kinase B Insulin IGF pathway protein kinase B signaling cascade 28 E 13 P00903 PDK1 2 Protein Protein kinase B Insulin IGF pathway protein kinase B signaling cascade 12 i Q
95. th complex vs mendelian disease evolutionary evidence for differences in molecular effects Proc Natl Acad Sci U S A 101 43 15398 403 10 2004 57 Index Activity Flow 26 Alternative ID 16 Analyzing gene list with expression values 48 Applet see PANTHER Pathway Applet attribute table 28 29 batch ID search 6 36 binomial distribution test 42 46 biological process 9 12 16 18 34 BioPAX 8 branch length 28 browse see prowler cellular component 9 12 16 18 35 closely related 38 coding SNP 53 collapse 28 Compare Gene List 42 Component see pathway molecule class component see pathway molecule class 13 21 22 Component Accession 13 Component Name 13 cSNP analysis tool 3 48 delete state 30 31 distantly related 39 Download 3 39 downstream 14 22 27 downstream path 27 ENSEMBL 6 Evidence 23 Evidence Code 23 expand 28 29 expected value 46 export 11 26 Family ID 11 family list page 11 Family Name 11 family subfamily detail page 18 file format 38 FTP see Download gene detail page 15 gene duplication 28 gene expression analysis tool 3 42 gene home page 6 Gene ID 9 16 gene list page 9 Gene Name 9 16 Gene Ontology 9 12 15 16 18 Gene Symbol 9 16 GO slim 3 9 12 14 15 18 30 34 heat map 51 hidden Markov model 7 18 HMM 3 7 18 length 18 score 38 scoring 3 7 38 home page 2 insert state 30 31 Interpro
96. tifiers in other databases for this gene or a protein product of the gene This is mapped based on the UniProt ID Mapping tool PANTHER GENE INFORMATION Gene ID ENSG00000135679 Protein ID Q00987 Gene Name E3 ubiquitin protein ligase Mdm2 Gene Symbol s MDM2 Organism Alternate Ids ENS 3148 Ensembl PRO ENSTO0000258148 Ensembl TRS MDM2 NP 0011388 Show All Figure 2 11 Gene information section of the gene detail page 2 PANTHER classification of the gene Figure 2 12 PANTHER family The family to which the gene belongs there is a phylogenetic tree for this family This link will bring up the PANTHER family page PANTHER subfamily The subfamily to which the gene belongs the subfamily is an notated on the phylogenetic tree This link will bring up the PANTHER subfamily page GO molecular function biological process cellular component These are Gene Ontology terms describing the function of the gene product PANTHER Protein Class This isa PANTHER Index terms describing protein classes Pathway This links to a diagram of the pathway s a gene product participates in CHAPTER 2 PANTHER AT A GLANCE 17 PANTHER CLASSIFICATION PANTHER Family MOM2 4 PANTHER Subfamily 3 UBIQUITIN PROTEIN LIGASE MDM2 PS3 BINDING PROTEIN MDM2 Le HE Tree MSA GO Molecular Function Catalytic activity 4 ligase activity L ubiquitin protein ligase activity GO Biological Process apoptosis 4 neg
97. tton Here is how the result page is read Figure 3 7 e PANTHER hit Only the top hit HMM is reported here The family or subfamily id and name of the top hit are displayed on the page Clicking the name will take you to the family subfamily detail page section 2 3 2 e PANTHER score The score of the PANTHER hit The green dots next to the score indicates how closely related the protein is to the model There are three categories closely related indicated by 3 greet dots if the score is better than E 23 very likely to be a correct functional assignment related indicated by 2 green dots if the score is better than E 11 but worse than E 23 molecular function likely to be the correct but biological process pathway less certain CHAPTER 3 PANTHER BASICS 39 distantly related indicated by 1 green dot if the score is better than E 3 but worse than E 11 protein is evolutionarily related but function may have diverged PANTHER HMM SEQUENCE SCORING RESULTS The top scoring HMM is reported along with the E value the number of expected false positive hits expected If the E value is less than 1e 3 no hits are reported PANTHER Hit APOLIPOPROTEIN E PTHR18976 SF2 HMM E value score 2 2e 155 eee C Sequence Domain seq f seq t hmm f hmm t score E value sequence 1 1 1 317 1 331 527 0 2 2e 155 Alignments of top scoring domains sequence domain 1 of 1 from 1 to 317 score 527 0 E 2 2e 155
98. within the family The longer the horizontal branch length the more distant the groups joined by those branches Vertical branch length is fixed for ease of viewing together with the information on the panel There are 4 types of nodes in the phylogenetic tree e Blue diamonds Subfamily nodes Subfamilies are usually colored to help distinguish between different subfamilies Aside from this the subfamily color does not have any special significance e Orange circles Gene duplication nodes e Green circles Speciation nodes e End of the horizontal branch Leaf nodes They usually point to the sequence IDs The following are a few things that you can do to these nodes 1 When mouse over these nodes the subfamily annotation information is displayed For the the subfamily node blue diamond it displays the subfamily id and name For all other node types it displays which subfamily it belongs to 2 When a node is clicked except for the leaf node it collapses e A collapsed subfamily view provides a high level view of the tree in which subfamilies are the most specific leaves of the tree The subfamily name given by curators appears in the sf name column of the Collapsed view The GO and PANTHER protein class terms are shown in the appropriate columns also e A collapsed non subfamily node is shown as a red circle The annotation node AN id is shown in the Protein id column and the common GO and protein class anno
99. ym 22 tool 9 training sequence 13 21 tree 3 tree home page 7 59 tree viewer 28 under representation 47 UniProt 6 upstream 14 22 27 upstream path 27 Wilcoxon Rank Sum test 48 50 51 workspace 3 9 11 40 organize 41 share 41

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