Systematic integration of millions of peptidoform evidences into Ensembl and other genome browsers
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There is an increasing demand for approaches to integrate proteomics data with other ‘omics’ data types, especially genomics. In fact, current resources to integrate proteomics in a genome context are insufficient for large-scale studies. To bring the genomics and proteomics results into coherence, novel resources must be developed that provide simple links across previously acquired datasets with minimal preprocessing and hassle. PRIDE (http://www.ebi.ac.uk/pride) and Ensembl (http://www.ensembl.org) are world-leading resources for proteomics and genomics data. We have developed a new resource and framework to enable a systematic integration of mass spectrometry (MS) based proteomics evidences into genome browsers. We automatically integrate every high-quality MS-based peptidoform reported in the PRIDE database, into genome coordinates through Ensembl, and other genome browsers.
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Systematic integration of millions of peptidoform evidences into Ensembl and other genome browsers
- 1. Yasset Perez-Riverol Ph.D PRIDE Team Systematic integration of millions of peptidoform evidences into Ensembl and other genome browsers.
- 2. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Proteogenomics • Provides a Trackhub in ENSEMBL for every ProteomeXchange COMPLETE submission in ENSEMBL. • Provides a global TrackHub in ENSEMBL for all PRIDE peptide evidences.
- 3. Meeting Sanger-EBI team EMBL-EBI, November 2017 PX Complete Trackhub in ENSEMBL PX Submission Tool PRIDE Archive 1 2 PRIDE submission Pipelines PRIDE Archive Web and API 3 TrackHub Registry 4 PX submission can be Partial or Complete: Partial Submission: RAW data, SEARCH Results and Peaks Lists. Complete Submission: RAW data, Result Files and Peak Lists, SEARCH Results. 1 Each PX submission can be search by: Title, Metadata, Description, Tissue, Taxonomy, PTMs. Peptide Sequence or Protein Identifier. 3 TrackHub Registry can search Tracks by: ShortLabel, LongLabel. OmicsType: Proteomics, Genomics, Transcriptomics. 4 PRIDE Submission Pipelines2 mzid Peak lists PX Complete Submission Assays mztab mgf lists PX Complete Submission Assays 5 Convert to mztab/mgf and filter evidences do not pass the reported mzid threshold. 5 Storage of the Project Metadata, Peptide Sequences, Protein identifiers in Solr and MongoDB. 6 6 Assay Peptide Pogo File PX Complete Submission Taxonomies Track TrackHub Generation Taxonomies TrackHub Registry
- 4. Meeting Sanger-EBI team EMBL-EBI, November 2017 Generation reliable peptide tables Current Filter options: • 1% FDR PSM level (Combine Results) • 1% FDR Peptide Level (Combine Results) Possible Filters (HPP): • > 8 AA • 1% FDR at transcript level (inference needed) Combine PSM Score: - Same Spectra, Peptide - Different Search Engine Combine Peptide Score: - Same Peptide - Different PSMs Experiment Peptide PSMs Quant <a href="http://www.ebi.ac.uk/pride/archive/assays/34642">Assay 34642</a> APPLLEGAPFR 1 1.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34644">Assay 34644</a> APPLLEGAPFR 1 1.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34642">Assay 34642</a> THTQDAVPLTLGQEFSGYVQQVQYAM(oxidation)VR 1 1.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34645">Assay 34645</a> KKQVM(oxidation)EK 1 1.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34645">Assay 34645</a> VGSGDTNNFPYLEK 2 2.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34645">Assay 34645</a> SLTYLSILR 3 3.000000 <a href="http://www.ebi.ac.uk/pride/archive/assays/34642">Assay 34642</a> LPFTPLSYIQGLSHR 8 8.000000 Audain et. al, J Proteomics 2017 A B DC E P1 P3P2 P4 PR1 JIG HF P5 PR1 Protein Inference Toolkit Protein Groups
- 5. Meeting Sanger-EBI team EMBL-EBI, November 2017 Mapping peptides to ENSEMBL: PoGo Schlaffner CN., Pirklbauer G, Bender A , Choudhary JS, PoGo: Jumping from Peptides to Genomic Loci, biorxiv (2016) For each .pogo file: • PTMs are standard to a common representation using PRIDE-Mod library. • Each Peptide reference to an Assay URL in PRIDE. • Each Pogo file is generated automatically by the PRIDE Pipeline. chr1 1314335 1314365 VLIPVFALGR 1000 - 1314335 1314335 0,0,0 1 30 0 chr1 1454464 1454488 ITVLEALR 1000 + 1454464 1454464 128,128,128 1 24 0 chr1 1456317 1456344 LFDWANTSR 1000 + 1456317 1456317 128,128,128 1 27 0 chr1 1459184 1459211 ATLNAFLYR 1000 + 1459184 1459184 128,128,128 1 27 0 chr1 1462609 1462633 LAQFDYGR 1000 + 1462609 1462609 128,128,128 1 24 0 chr1 1485135 1485159 ITVLEALR 1000 + 1485135 1485135 128,128,128 1 24 0 chr1 1486636 1486663 LFDWANTSR 1000 + 1486636 1486636 128,128,128 1 27 0 chr1 1490572 1490596 LAQFDYGR 1000 + 1490572 1490572 128,128,128 1 24 0 chr1 1522863 1522887 ITVLEALR 1000 + 1522863 1522863 128,128,128 1 24 0 Challenge in the Future: • Bed information can be extended with more information about the transcript reliability. • Peptide uniqueness • Reliability score. • Native bigBed should be provided to remove the customization of new pipelines, etc. • What to do with the unmapped peptides (which are long lists.) • Maintainability.
- 6. Meeting Sanger-EBI team EMBL-EBI, November 2017 Getting there.. Taxonomy COUNT Human 876 Mouse 252 Arabidopsis thaliana 127 Rat 68 Yeast 62 E.Coli 38 Bos taurus 32 Drosophila melanogaster 21 zebrafish 20 Zeaa Mays 15 Candida albicans 12 Gallus gallus 12 Caenorhabditis elegans 10 Horse 10 Synechocystis sp. PCC 9 Glycine max 9 Complete Submission PRIDE Taxonomy COUNT Human 74 Mouse Arabidopsis thaliana Rat 4 Yeast E.Coli Bos taurus Drosophila melanogaster zebrafish Zeaa Mays Candida albicans Gallus gallus Caenorhabditis elegans Horse Synechocystis sp. PCC Glycine max TrackHub Creation
- 7. Meeting Sanger-EBI team EMBL-EBI, November 2017 TrackHub creation and Publication Track TrackHub Creation Taxonomies Pogo Files Pogo Files Taxonomies TrackHub Registry trackhub- parameters.json 1 2 TrackHub 3 TrackHub Registry 4 TrackHub Creator: Python Framework. Interact with ENSEMBL API to retrieve the latest ENSEMBL release and assembly version (for all species supported by PoGo. Download the corresponding GTF and FASTA (Protein File) for each Species supported by PoGo. Using Pogo source code, compile the Pogo version and generate the bed/bigbed files. Create/Update the TrackHub for an specific PX submission. A new library has been developed to interact with ENSEMBL trackhub registry (https://github.com/PRIDE-Utilities/track- hub-registrator) https://github.com/Proteogenomics/trackhub- creator 1 2 3 4
- 8. Meeting Sanger-EBI team EMBL-EBI, November 2017 Trackhub creator • Download ENSEMBL FASTA Protein file and GTF for an specific taxonomy. Detect the taxonomy in PRIDE and automatically query ENSEMBL API and Download the data from it. • Run PoGo and generate the mapping files: bed files for Peptides and Modified Peptides (On going, We will automatically generate the 1 and 2 gaps files). • Generate the TrackHub with an specific structure (Next session.). • Publish the Trackhub in ENSEMBL Registry (https://www.trackhubregistry.org/search). Note: We have faced millions of problems: 1- ENSEMBL and UCSC support only UCSC notations for almost everything: genome hg38 trackDb hg38/trackDb.txt Chromosomes should be chr1, chr2.. Etc. Chromosomes sizes are programmatically available. 2- Taxonomies where not supported on PoGo. We have moved to a better list now with 19 Taxonomies. 3- Modifications has been a nightmare between UNIMOD and PoGo. http://ftp.pride.ebi.ac.uk/pride/data/proteogenomics/latest/archive/
- 9. Meeting Sanger-EBI team EMBL-EBI, November 2017 Getting there.. http://genome.ucsc.edu/cgi- bin/hgTracks?db=mm10&lastVirtModeType=default&lastVirtModeExtraState=&virtModeType=default&virtMode= 0&nonVirtPosition=&position=chr12%3A20021505- 100107519&hgsid=644445905_BScrPMQymPnGtl9O7jeSv1jS5Rm0
- 10. Meeting Sanger-EBI team EMBL-EBI, November 2017 Getting there..
- 11. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Proteogenomics • Provides a Trackhub in ENSEMBL for every ProteomeXchange COMPLETE submission in ENSEMBL. • Provides a global TrackHub in ENSEMBL for all PRIDE peptide evidences.
- 12. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Data Growth
- 13. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Cluster Approach “When merging large number of datasets coming from different groups/bioinformatics workflows the consensus spectra should be generated to evaluate the accuracy of each independent dataset." We know results depend on: Database, Search engine Settings, bioinformatics pipeline, etc. Assay-24261 Assay-3394 Peptide Mascot Score: 54.06 Peptide Mascot Score: 32.86 GYTFSTTAER: 3 PSMs GYSFTTTAER: 13918 (143)
- 14. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Cluster Pipeline PX Complete . . n Hadoop Cluster PRIDE Archive Import PX successfully converted New Peptide/PTMs Number of Identified and non-Identified Spectra QC QC Number of new clusters PRIDE Cluster score distribution Number of clusters by modification mgf (Annotated spectra) Clustering Files QC Number of Peptides Number of new Peptides Number of PTMs Number of New PTMs Peptide Tables (Pogo File) Peptide Export Taxonomies Track TrackHub Generation Taxonomies TrackHub Registry
- 15. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE Cluster Peptide Evidences. Human MouseArabidopsis The Peptide Evidence Files from PRIDE Cluster collapsed all the peptide evidences from reliable clusters: ftp://ftp.pride.ebi.ac.uk/pride/data/cluster/peptide- results/2015-04/
- 16. Meeting Sanger-EBI team EMBL-EBI, November 2017 All trackhubs already published Note: Some challenges ahead: 1- Taxonomies like Saccharomyces cerevisiae are not supported in PoGo. 2- Scaffold are not supported in Pogo with the UCSC notation. The same problem that we face with chromosome 1, 2, .. To chr1 chr2, etc can’t be done with scaffold. 3- More evidences are needed.
- 17. Meeting Sanger-EBI team EMBL-EBI, November 2017 Getting there … (PepBed package) • Black (all identified peptides). • Cyan (oxidation) • Orange (acetyl) • Red (phospho)
- 18. Meeting Sanger-EBI team EMBL-EBI, November 2017 Getting there … (Comparions with APRIS Peptidome) APRIS Peptidome: • The eight studies covered a huge range of tissues and cell types: • The peptides from the PeptideAtlas database cover 51 different tissues, cell types, and developmental stages (2016). • Geiger study interrogated 11 different cell types. • NIST database. • The Kim and Wilhelm analyses peptides were generated from 30 and 35 distinct tissues types (51 tissues in total).
- 19. Meeting Sanger-EBI team EMBL-EBI, November 2017 PRIDE team. Manuel Bernal-Llinares (track-hub creator) Tobias Ternent (pride pipelines) Johannes Griss (pride cluster pipelines) Sanger Team Christoph Schlaffner (pogo tool) Jyoti Choudhary (PI) Juan A. Vizcaino (PI) Alessandro Vullo (trackhub registry) ENSEMBLTeam