Yale Pseudogene Analysis as part of GENCODE Project Sanger Center  2009.01.20, 11:20-11:40 Mark B Gerstein Yale   (c) Mark...
Overview <ul><li>Outline </li></ul><ul><ul><li>Flow </li></ul></ul><ul><ul><li>Additional Pseudogene Annotation  </li></ul...
Overall Flow: Pipeline Runs, Coherent Sets, Annotation, Transfer to Sanger  <ul><li>Overall Approach </li></ul><ul><ul><li...
Additional Annotation on Pseudogenes: Consistent Labeling (Ontology)
Gene duplicated_from mutated_from retrotranscribed_from Protein family contained_in has_pseudogene_family mRNA Protein tra...
Pseudogene Duplicated Unitary Transcribed Regulatory Processed Classified Type Semi Processed Duplicated Processed Polymor...
Collections to provide further annotation OTTHUMG00000000445 HAVANA ENSP00000384933.frag.301873 BC067222.1-28 urn:lsid:pse...
Additional Annotation on Pseudogenes: Families & Histories
Flat Files Table Browser tables.pseudogene.org DAS UCSC Genome Browser <ul><li>12 eukaryotic species </li></ul><ul><ul><li...
Pseudofam Database <ul><li>Data Sources </li></ul><ul><ul><li>Ensembl, Pfam, BioMart </li></ul></ul><ul><ul><li>Pseudopipe...
Pseudofam Construction <ul><li>Data Generation </li></ul><ul><ul><li>Identify pseudogenes by proteins </li></ul></ul><ul><...
Pseudofam Statistics: Enrichment of pseudogenes within a family (&quot;Living vs Dead&quot;) Total (10 Eukaryotes) Human [...
Pseudogene Set #2:  RP pseudogenes
Human Ribosomal Proteins (RP) [Zhang  et al .  Genome Research ,  2002] 79 Functional RP genes Nakao A, Yoshihama M, Kenmo...
Human Ribosomal Proteins (RP) ~ 2000 RP   genes [Zhang  et al .  Genome Research ,  2002] 79 Functional RP genes
Number of RP pseudogenes (identified by pipeline) RP pseudogenes constitute the largest family of pseudogenes. Almost all ...
Number of each type of human ribosomal protein processed pseudogenes appears unrelated to expression level or to number in...
Using Synteny to Improve Annotation of RP pgenes  Synteny derived based on local gene orthology [Balasubramanian et al.,  ...
Syntenic proc pseudogenes [Balasubramanian et al.,  Genome Biol.  ('09)] Species1- Species2 Number of syntenic pgenes Huma...
A human-mouse syntenic pgene, likely to be coding [Balasubramanian et al.,  Genome Biol.  ('09)] Nakao A, Yoshihama M, Ken...
Set #3: Transcribed Pseudogenes Annotated Earlier
Harrison '05 set of  Transcribed Pseudogenes <ul><li>233  Transcribed from ~8000 Processed Pseudogenes  </li></ul><ul><li>...
Set #4: Strong Consensus Pseudogenes
Consensus Pseudogenes <ul><li>Yale & UCSC agreement  </li></ul><ul><li>Yale, UCSC & HAVANA agreement </li></ul><ul><li>HAV...
<ul><li>Initially, 50 nucleotide overlap </li></ul><ul><li>Why so low? </li></ul><ul><li>What about percentage overlap? </...
 
 
 
High Quality  Overlaps?
 
gencode.gersteinlab.org/consensus <ul><li>Django-based </li></ul><ul><li>Filter by genomic coordinates, source  </li></ul>...
Set #5: SD pseudogenes
Segmental Duplications (SDs) <ul><li>SDs are regions of the genome with    90% sequence identity and    1kb in length </...
Pseudogene families and Segmental Duplications (SDs) <ul><li>SDs comprise ~5-6% of the human genome but contain ~17.8% gen...
Summary <ul><li>Sets </li></ul><ul><ul><li>RP, transcribed ‘05, consensus, SD....  </li></ul></ul><ul><li>Additional Annot...
Credits <ul><li>Yale:  S Balasubramanian, P Cayting, H Lam,  Y Liu, G Fang, N Carriero, R Robilotto, E Khurana </li></ul><...
Extra
Collections <ul><li>Combination of multiple sets </li></ul><ul><ul><li>Human50, UCSC, GAPDH, Transcribed, Unitary, etc. </...
Pseudofam & Pseudogene.org Integration PseudoPipe svn annotations  via shared ID
gencode.gersteinlab.org/consensus
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http://lectures.gersteinlab.org/ppt/Gencode-winter08-20090121-pseudogenes/Gencode-winter08-20090121-pseudogenes

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Yale Pseudogene Analysis as part of GENCODE Project
Sanger Center 2009.01.20, 11:20-11:40
Date Given: 01/21/2009

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http://lectures.gersteinlab.org/ppt/Gencode-winter08-20090121-pseudogenes/Gencode-winter08-20090121-pseudogenes

  1. 1. Yale Pseudogene Analysis as part of GENCODE Project Sanger Center 2009.01.20, 11:20-11:40 Mark B Gerstein Yale (c) Mark Gerstein, 2002, Yale, bioinfo.mbb.yale.edu Illustration from Gerstein & Zheng (2006). Sci Am.
  2. 2. Overview <ul><li>Outline </li></ul><ul><ul><li>Flow </li></ul></ul><ul><ul><li>Additional Pseudogene Annotation </li></ul></ul><ul><ul><li>Pseudogene Sets </li></ul></ul><ul><li>Regular conference calls </li></ul><ul><ul><li>Sanger / Havana: Jenn, Adam </li></ul></ul><ul><ul><li>UCSC: Rachel, Mark </li></ul></ul><ul><ul><li>Discuss difficult pseudogene cases, pipeline improvements and additional pseudogene annotation </li></ul></ul><ul><li>Questions for you: </li></ul><ul><ul><li>consis. Labels ? </li></ul></ul><ul><ul><li>Pfam/ensembl usage ? </li></ul></ul><ul><ul><li>trans. set ? </li></ul></ul><ul><ul><li>consensus set ? </li></ul></ul>
  3. 3. Overall Flow: Pipeline Runs, Coherent Sets, Annotation, Transfer to Sanger <ul><li>Overall Approach </li></ul><ul><ul><li>Overall Pipeline runs at Yale and UCSC, yielding raw pseudogenes </li></ul></ul><ul><ul><li>Extraction of coherent subsets for further analysis and annotation </li></ul></ul><ul><ul><li>Passing to Sanger for detailed manual analysis and curation </li></ul></ul><ul><ul><li>Incorporation into final GENCODE annotation </li></ul></ul><ul><ul><li>Pipeline modification </li></ul></ul><ul><li>Chronology of Sets </li></ul><ul><ul><li>Unitary pseudogenes </li></ul></ul><ul><ul><li>Ribosomal Protein pseudogenes </li></ul></ul><ul><ul><ul><li>(Hard then easy) </li></ul></ul></ul><ul><ul><li>Transcribed pseudogenes annotated earlier </li></ul></ul><ul><ul><li>Strong overlap consensus pseudogenes </li></ul></ul><ul><ul><li>Pseudogenes associated with SDs </li></ul></ul><ul><ul><li>GAPDH pseudogenes </li></ul></ul>
  4. 4. Additional Annotation on Pseudogenes: Consistent Labeling (Ontology)
  5. 5. Gene duplicated_from mutated_from retrotranscribed_from Protein family contained_in has_pseudogene_family mRNA Protein transcribed_into translated_into Pseudogene family type_of Sequence Region Segmental Duplication Syntenic Region etc is_a contained_in contained_in has_parent_protein derives_from Domain Ontology Pseudogene Upper Ontology [Lam et al., NAR DB Issue (in press, '09)] Pseudogene Duplicated Unitary Transcribed Regulatory Processed … …
  6. 6. Pseudogene Duplicated Unitary Transcribed Regulatory Processed Classified Type Semi Processed Duplicated Processed Polymorphic Feature Evidence Sequence Homology Intra-Genome Homology Cross-Genome Homology Recognition Feature Disablement Premature Stop Codon Regulatory Element Lost Frameshift Pseudo-PolyA Tail Nucleus Mitochondria Origin has a is a Domain Ontology *Unprocessed Proposed HAVANA [Lam et al., NAR DB Issue (in press, '09)]
  7. 7. Collections to provide further annotation OTTHUMG00000000445 HAVANA ENSP00000384933.frag.301873 BC067222.1-28 urn:lsid:pseudogene.org:9606.Pseudogene:27570 urn:lsid:pseudogene.org:9606.Pseudogene:27579 urn:lsid:pseudogene.org:9606.Pseudogene:29 Human Pseudogenes Yale UCSC Ribosomal Protein Transcribed
  8. 8. Additional Annotation on Pseudogenes: Families & Histories
  9. 9. Flat Files Table Browser tables.pseudogene.org DAS UCSC Genome Browser <ul><li>12 eukaryotic species </li></ul><ul><ul><li>Human, mouse, rat, chimp… </li></ul></ul><ul><ul><li>100,052 pseudogenes </li></ul></ul><ul><li>64 prokaryotic species </li></ul><ul><ul><li>6,412 pseudogenes </li></ul></ul><ul><li>28,237 human pseudogenes </li></ul><ul><ul><li>13+ unique human sets </li></ul></ul>SVN [Karro et al., NAR ('07)] single pgene query
  10. 10. Pseudofam Database <ul><li>Data Sources </li></ul><ul><ul><li>Ensembl, Pfam, BioMart </li></ul></ul><ul><ul><li>Pseudopipe </li></ul></ul><ul><li>Highlighted Features </li></ul><ul><ul><li>Browse families </li></ul></ul><ul><ul><ul><li>Statistics </li></ul></ul></ul><ul><ul><ul><ul><li>Enrichment </li></ul></ul></ul></ul><ul><ul><ul><ul><li>P-value, etc </li></ul></ul></ul></ul><ul><ul><li>Search families </li></ul></ul><ul><ul><ul><li>Pfam ID/Acc </li></ul></ul></ul><ul><ul><ul><li>Ensembl ID </li></ul></ul></ul><ul><ul><ul><li>Pseudogene ID </li></ul></ul></ul><ul><ul><li>Correlate families </li></ul></ul><ul><ul><ul><li>Genes </li></ul></ul></ul><ul><ul><ul><li>Pseudogenes </li></ul></ul></ul><ul><ul><ul><li>Parents </li></ul></ul></ul>[Lam et al., NAR DB Issue (in press, '09)]
  11. 11. Pseudofam Construction <ul><li>Data Generation </li></ul><ul><ul><li>Identify pseudogenes by proteins </li></ul></ul><ul><ul><li>Map parent proteins to protein families </li></ul></ul><ul><ul><li>Assign pseudogenes to their parent families </li></ul></ul><ul><ul><li>Align the pseudogenes in pseudogene families </li></ul></ul><ul><ul><li>Calculate the key statistics and organize the data into database </li></ul></ul><ul><li>Alignment </li></ul><ul><ul><li>Align pseudogene to parent </li></ul></ul><ul><ul><li>Transfer alignment from Pfam </li></ul></ul><ul><ul><li>Combine and adjust the alignments to build the pseudofam alignment </li></ul></ul>[Lam et al., NAR DB Issue (in press, '09)]
  12. 12. Pseudofam Statistics: Enrichment of pseudogenes within a family (&quot;Living vs Dead&quot;) Total (10 Eukaryotes) Human [Lam et al., NAR DB Issue (in press, '09)]
  13. 13. Pseudogene Set #2: RP pseudogenes
  14. 14. Human Ribosomal Proteins (RP) [Zhang et al . Genome Research , 2002] 79 Functional RP genes Nakao A, Yoshihama M, Kenmochi N: RPG: the Ribosomal Protein Gene database. Nucleic Acids Res 2004, 32:D168-170.
  15. 15. Human Ribosomal Proteins (RP) ~ 2000 RP  genes [Zhang et al . Genome Research , 2002] 79 Functional RP genes
  16. 16. Number of RP pseudogenes (identified by pipeline) RP pseudogenes constitute the largest family of pseudogenes. Almost all are processed: There are ~90 clearly duplicated ones in the human genome [Balasubramanian et al., Genome Biol. ('09)] Organism Processed Fragments Low confidence Human 1822 218 212 Chimp 1462 219 160 Mouse 2092 326 413 Rat 2848 343 450
  17. 17. Number of each type of human ribosomal protein processed pseudogenes appears unrelated to expression level or to number in mouse [Balasubramanian et al., Genome Biol. ('09)]
  18. 18. Using Synteny to Improve Annotation of RP pgenes Synteny derived based on local gene orthology [Balasubramanian et al., Genome Biol. ('09)]
  19. 19. Syntenic proc pseudogenes [Balasubramanian et al., Genome Biol. ('09)] Species1- Species2 Number of syntenic pgenes Human-chimp 1282 Human-mouse 6 Human-rat 11 Rat-mouse 394
  20. 20. A human-mouse syntenic pgene, likely to be coding [Balasubramanian et al., Genome Biol. ('09)] Nakao A, Yoshihama M, Kenmochi N: RPG: the Ribosomal Protein Gene database. Nucleic Acids Res 2004, 32:D168-170.
  21. 21. Set #3: Transcribed Pseudogenes Annotated Earlier
  22. 22. Harrison '05 set of Transcribed Pseudogenes <ul><li>233 Transcribed from ~8000 Processed Pseudogenes </li></ul><ul><li>Evidence for Transcription </li></ul><ul><ul><li>8% Refseq mRNAs </li></ul></ul><ul><ul><li>32% Unigene consensus sequences </li></ul></ul><ul><ul><li>72% dbEST expressed sequence tags </li></ul></ul><ul><ul><li>32% Oligonucleotide microarray data (extra support) </li></ul></ul><ul><li>Highly decayed </li></ul><ul><ul><li>Fraction with Ka/Ks ≥ 0.5 is 54% </li></ul></ul>Harrison et al. (2005) NAR
  23. 23. Set #4: Strong Consensus Pseudogenes
  24. 24. Consensus Pseudogenes <ul><li>Yale & UCSC agreement </li></ul><ul><li>Yale, UCSC & HAVANA agreement </li></ul><ul><li>HAVANA disagreement </li></ul>
  25. 25. <ul><li>Initially, 50 nucleotide overlap </li></ul><ul><li>Why so low? </li></ul><ul><li>What about percentage overlap? </li></ul>Overlap
  26. 29. High Quality Overlaps?
  27. 31. gencode.gersteinlab.org/consensus <ul><li>Django-based </li></ul><ul><li>Filter by genomic coordinates, source </li></ul><ul><li>Yale & UCSC Consensus track </li></ul><ul><li>Flagged pseudogenes </li></ul><ul><li>Verification status </li></ul>
  28. 32. Set #5: SD pseudogenes
  29. 33. Segmental Duplications (SDs) <ul><li>SDs are regions of the genome with  90% sequence identity and  1kb in length </li></ul><ul><li>Clues about pseudogene formation from pseudogenes located in SDs, for example, annotation of “duplicated-processed” pseudogenes </li></ul>
  30. 34. Pseudogene families and Segmental Duplications (SDs) <ul><li>SDs comprise ~5-6% of the human genome but contain ~17.8% genes, 45.7% duplicated pgenes and 21.6% processed pgenes </li></ul><ul><li>Relative values of correlation coefficients in the plots above consistent with the observation that SDs contain more pgenes than parent genes </li></ul>[Lam et al., NAR DB Issue (in press, '09)]
  31. 35. Summary <ul><li>Sets </li></ul><ul><ul><li>RP, transcribed ‘05, consensus, SD.... </li></ul></ul><ul><li>Additional Annotation </li></ul><ul><ul><li>families & labels </li></ul></ul>
  32. 36. Credits <ul><li>Yale: S Balasubramanian, P Cayting, H Lam, Y Liu, G Fang, N Carriero, R Robilotto, E Khurana </li></ul><ul><li>Alums: D Zheng, P Harrison, Z Zhang </li></ul><ul><li>Sanger: A Frankish, J Harrow </li></ul><ul><li>UCSC: R Harte, M Diekhans </li></ul>
  33. 37. Extra
  34. 38. Collections <ul><li>Combination of multiple sets </li></ul><ul><ul><li>Human50, UCSC, GAPDH, Transcribed, Unitary, etc. </li></ul></ul><ul><li>Also can have attributes </li></ul><ul><li>View by set or as a whole </li></ul>
  35. 39. Pseudofam & Pseudogene.org Integration PseudoPipe svn annotations via shared ID
  36. 40. gencode.gersteinlab.org/consensus

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