pride and proteomexchange: training webinar

PRIDE and ProteomeXchange: Training webinar

Dr. Juan Antonio Vizcaíno

PRIDE Group CoordinatorProteomics Services TeamEMBL-EBIHinxton, Cambridge, UKjuan@ebi.ac.uk

Juan A. Vizcaínojuan@ebi.ac.uk

Training webinar25 November 2015

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Data resources at EMBL-EBIGenes, genomes & variation

RNA CentralArrayExpress

Expression AtlasMetabolights

InterPro Pfam UniProt

ChEMBL ChEBI

Molecular structuresProtein Data Bank in EuropeElectron Microscopy Data Bank

European Nucleotide ArchiveEuropean Variation ArchiveEuropean Genome-phenome Archive

Gene, protein & metabolite expression

Protein sequences, families & motifs

Chemical biologyReactions, interactions & pathways

IntActReactome

MetaboLights

SystemsBioModels Enzyme Portal BioSamples

Ensembl Ensembl Genomes

GWAS CatalogMetagenomics portal

Europe PubMed CentralGene OntologyExperimental Factor Ontology

Literature & ontologies

Data resources at EMBL-EBIGenes, genomes & variation

RNA CentralArrayExpress

Expression AtlasMetabolights

InterPro Pfam UniProt

ChEMBL ChEBI

Molecular structuresProtein Data Bank in EuropeElectron Microscopy Data Bank

European Nucleotide ArchiveEuropean Variation ArchiveEuropean Genome-phenome Archive

Gene, protein & metabolite expression

Protein sequences, families & motifs

Chemical biologyReactions, interactions & pathways

IntActReactome

MetaboLights

SystemsBioModels Enzyme Portal BioSamples

Ensembl Ensembl Genomes

GWAS CatalogMetagenomics portal

Europe PubMed CentralGene OntologyExperimental Factor Ontology

Literature & ontologies

• PRIDE Archive (in the context of ProteomeXchange and the PSI standards)

• How to submit data to PRIDE: PRIDE tools

• How to access data in PRIDE Archive

• A sneak peak to other PRIDE resources

Overview

Training webinar25 November 2015 7

Mass Spectrometry (MS)-based proteomics• Many different workflows.

• Discovery mode:• Bottom-up proteomics

• Data dependent acquisition• Data independent acquisition

• Top down proteomics

• Targeted mode:• SRM (Selected Reaction Monitoring)

Training webinar25 November 2015 8

Mass Spectrometry (MS)-based proteomics• Many different workflows.

• Discovery mode:• Bottom-up proteomics

• Data dependent acquisition• Data independent acquisition

• Top down proteomics

• Targeted mode:• SRM (Selected Reaction Monitoring)

MS proteomics: tandem MS (bottom-up)

MS/MS matching identifies peptides, not proteins.

Proteins are inferred from the peptide sequences.

• PRIDE stores mass spectrometry (MS)-based proteomics data:

• Peptide and protein expression data (identification and quantification)

• Post-translational modifications• Mass spectra (raw data and peak

lists)• Technical and biological metadata• Any other related information

• Full support for tandem MS approaches

PRIDE (PRoteomics IDEntifications) database

http://www.ebi.ac.uk/pride/archive Martens et al., Proteomics, 2005Vizcaíno et al., NAR, 2013

PRIDE Mission

• To archive all types of proteomics mass spectrometry data for the purpose of supporting reproducible research, allowing the application of quality control metrics and enabling the reuse of these data by other researchers.

• To integrate MS-based data in a protein-centric manner to provide information on protein variants, modifications, and expression.

• To provide mass spectrometry based expression data to the Expression Atlas.

PRIDE Mission

• To archive all types of proteomics mass spectrometry data for the purpose of supporting reproducible research, allowing the application of quality control metrics and enabling the reuse of these data by other researchers.

• To integrate MS-based data in a protein-centric manner to provide information on protein variants, modifications, and expression.

• To provide mass spectrometry based expression data to the Expression Atlas.

What is a proteomics publication in 2015?• Proteomics studies generate potentially large amounts of

data and results.

• Ideally, a proteomics publication needs to:• Summarize the results of the study• Provide supporting information for reliability of any

results reported

• Information in a publication:• Manuscript• Supplementary material• Associated data submitted to a public repository

Journal Submission Recommendations• Journal guidelines recommend submission to proteomics repositories:

Proteomics Nature Biotechnology Nature Methods Molecular and Cellular Proteomics

• Funding agencies are enforcing public deposition of data to maximize the value of the funds provided.

PRIDE: Source of MS proteomics data

• PRIDE Archive already provides or will soon provide MS proteomics data to other EMBL-EBI resources such as UniProt, Ensembl and the EBI Expression Atlas.

http://www.ebi.ac.uk/pride/archive

Data content in PRIDE Archive• Dataset submission driven resource.

• PRIDE is organised in datasets (group of assays).

• An assay represents one MS run (in most cases).

• No data reprocessing at present. PRIDE aims to represent the author’s view on the data.

• Main supported formats: PRIDE XML and mzIdentML.

• Raw data is also now stored.

ProteomeXchange Consortium•Goal: Development of a framework to allow

standard data submission and dissemination pipelines between the main existing proteomics repositories.

•Includes PeptideAtlas (ISB, Seattle), PRIDE (Cambridge, UK) and (very recently) MassIVE (UCSD, San Diego).

•Common identifier space (PXD identifiers)

•Two supported data workflows: MS/MS and SRM.

•Main objective: Make life easier for researchers

http://www.proteomexchange.org

ProteomeCentral

Metadata / Manuscript

Raw Data*

Results

Journals

UniProt/neXtProt

Peptide Atlas

Other DBs

Receiving repositories

PASSEL (SRM data)

PRIDE (MS/MS data)

Other DBs

Researcher’s results

Reprocessed results

Raw data*

Metadata

MassIVE (MS/MS data)

Vizcaíno et al., Nat Biotechnol, 2014

ProteomeXchange data workflow

Overview

ProteomeCentral

Raw Data*

Results

Journals

UniProt/neXtProt

Peptide Atlas

Other DBs

PASSEL (SRM data)

PRIDE (MS/MS data)

Other DBs

Reprocessed results

Raw data*

Metadata

PX Data workflow for MS/MS data1. Mass spectrometer output files: raw data (binary files) or

peak list spectra in a standardized format (mzML, mzXML).

2. Result files:

a. Complete submissions: Result files can be converted to PRIDE XML or the mzIdentML data standard.

b. Partial submissions: For workflows not yet supported by PRIDE, search engine output files will be stored and provided in their original form.

3. Metadata: Sufficiently detailed description of sample origin, workflow, instrumentation, submitter.

4. Other files: Optional files:a. QUANT: Quantification related results e. FASTAb. PEAK: Peak list files f. SP_LIBRARYc. GEL: Gel imagesd. OTHER: Any other file type

Published

RawFiles

Other files

Complete

Partial

Complete vs Partial submissions: processed resultsFor complete submissions, it is possible to connect the spectra with the identification

processed results and they can be visualized.

PX Data workflow for MS/MS data1. Mass spectrometer output files: raw data (binary files) or

peak list spectra in a standardized format (mzML, mzXML).

2. Result files:

a. Complete submissions: Result files can be converted to PRIDE XML or the mzIdentML data standard.

b. Partial submissions: For workflows not yet supported by PRIDE, search engine output files will be stored and provided in their original form.

3. Metadata: Sufficiently detailed description of sample origin, workflow, instrumentation, submitter.

4. Other files: Optional files (the list can be extended):a. QUANT: Quantification related results e. FASTAb. PEAK: Peak list files f. SP_LIBRARYc. GEL: Gel imagesd. OTHER: Any other file type

Published

RawFiles

Other files

PRIDE Components: Submission Process

PRIDE Converter 2

PRIDE Inspector PX Submission Tool

mzIdentML

PRIDE XML1

Search output files

Spectra files

Original data files ‘RESULT’ file generation Final ‘RESULT’ file

PRIDE XML

‘RESULT’

Before: only file conversion to PRIDE XML

File conversion

PRIDE Converter

Other tools, e.g. hEIDI

PX Data workflow for MS/MS dataSearch Engine

Results + MS files

PRIDE Converter 2

PRIDE XML

Coté & Griss et al., MCP, 2012

Other tools available:

- PRIDE Converter- PLGS (Waters)- Proteios- EasyProt- hEIDI- OmicsHub (Integromics)- PeptideShaker (Compomics)

PRIDE Converter 2

https://github.com/PRIDE-Toolsuite/pride-converter-2

- ‘Bulk’ conversion possible: Command Line mode- Virtually no limit in file sizes.

Tools ‘RESULT’ file generation Final ‘RESULT’ file

mzIdentML ‘RESULT’

Now: native file export to mzIdentML

Spectra files

(mzML, mzXML, mzData,

mgf, pkl,

ms2, dta, apl)

Mascot

ProteinPilot

Scaffold

Others

Native File export

Complete submissionsSearch Engine

Results + MS files

Search engines

mzIdentML

- Mascot- MSGF+- MyriMatch and related tools from D. Tabb’s

lab- OpenMS- PEAKS- PeptideShaker- ProCon (ProteomeDiscoverer, Sequest)- Scaffold- TPP via the idConvert tool (ProteoWizard)- ProteinPilot (from version 5.0)- X!Tandem native conversion (Beta,

PILEDRIVER)- Others: library for X!Tandem conversion, lab

internal pipelines, …- Crux

An increasing number of tools support export to mzIdentML 1.1

- Referenced spectral files need to be submitted as well (all open formats are supported).

Updated list: http://www.psidev.info/tools-implementing-mzIdentML#.

PRIDE Converter 2

mzIdentML

PRIDE XML

PRIDE Inspector Toolsuite

Wang et al., Nat. Biotechnology, 2012Perez-Riverol et al., MCP, 2016, in press

PRIDE Inspector

PRIDE Inspector Toolsuite supports:

- PRIDE XML- mzIdentML + all types of spectra files- mzML- mzTab identification and Quantification +

all types of spectra files

https://github.com/PRIDE-Toolsuite/

New visualisation functionality for Protein Groups

PRIDE Inspector ToolsuitePRIDE Inspector Toolsuite

Private review of files submitted to PRIDE https://github.com/PRIDE-Toolsuite/

PRIDE Converter 2

mzIdentML

PRIDE XML

•Capture the mappings between the different types of files.

•Make the file upload process straightforward to the submitter (It transfers all the files using Aspera or FTP).

PX submission tool

Published

Other files

http://www.proteomexchange.org/submission

PXsubmission

•Command line alternative: Using the Aspera file transfer protocol.

PX submission tool: step by step

PX submission tool: screenshots

Manuscript published detailing the process

Ternent et al., Proteomics, 2014http://www.proteomexchange.org/submission

Example dataset:PXD000764

- Title: “Discovery of new CSF biomarkers for meningitis in children”- 12 runs: 4 controls and 8 infected samples- Identification and quantification data

PRIDE Archive submitted datasets up until 1st November, 2015

• 1,259 submitted datasets by November 1st • 923 submitted datasets in 2014• In the last 6 months, 155 submitted datasets per month• Size: ~ 160 TB.

PRIDE: Size comparison with other EBI resources (May 2015)

2004 2006 2008 2010 2012 2014 20161E+07

1E+17Data accumulation by resource

Metabo-lites

ENA (less AE)

Chart generated by Guy Cochrane

Overview

Data access to PRIDE Archive• Look for particular datasets of interest:

• For data reuse: which particular proteins and peptides (including PTMs) have been detected.

• Data reinterpretation or re-analysis.

• Validation of the experimental results reported.

• Specific use cases for proteomics: spectral libraries, fragmentation models, SRM transitions,…

RSS feed for public datasets

http://groups.google.com/group/proteomexchange/feed/rss_v2_0_msgs.xml

Ways to access data in PRIDE Archive

• PRIDE web interface

• File repository

• REST web service

• PRIDE Inspector tool

PRIDE Archive web interface

PRIDE Archive web interface (2)

ProteomeCentral

Raw Data*

Results

Journals

UniProt/neXtProt

Peptide Atlas

Other DBs

PASSEL (SRM data)

PRIDE (MS/MS data)

Other DBs

Reprocessed results

Raw data*

Metadata

ProteomeCentral: Portal for all PX datasets

http://proteomecentral.proteomexchange.org/cgi/GetDataset

Overview

2015 overview of PRIDE resources

PRIDE Proteomes and PRIDE Cluster• Provide an aggregated and QC filtered peptide-

centric and protein centric view on PRIDE Archive data. http://www.ebi.ac.uk/pride/cluster/http://wwwdev.ebi.ac.uk/pride/proteomes/

• Main characteristics of PRIDE Archive and ProteomeXchange (PX)

• PX/PRIDE submission workflow for MS/MS data• PRIDE Inspector• PX submission tool

• PRIDE/ProteomeXchange has become the de facto standard for data submission and data availability in proteomics

Conclusions

Do you want to know a bit more…?

http://www.slideshare.net/JuanAntonioVizcaino

Aknowledgements: PeopleAttila CsordasTobias TernentNoemi del ToroGerhard Mayer (Bochum, de.NBI)

Johannes GrissYasset Perez-Riverol

Henning Hermjakob

Former team members: Rui Wang, Florian Reisinger and Jose A. Dianes

Acknowledgements: The PRIDE Team

• 9 December – UniProt website updates• 16 December – Ensembl release 83

All webinars @ 4:00pm GMT time unless statedFor details see: http://www.ebi.ac.uk/training/webinars

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