Human Protein Reference Database (HPRD—http://www.hprd.org/), initially described in 2003, is a database of curated proteomic information pertaining to human proteins. We have recently added a number of new features in HPRD. These include PhosphoMotif Finder, which allows users to find the presence of over 320 experimentally verified phosphorylation motifs in proteins of interest. Another new feature is a protein distributed annotation system—Human Proteinpedia (http://www.humanproteinpedia.org/)—through which laboratories can submit their data, which is mapped onto protein entries in HPRD. Over 75 laboratories involved in proteomics research have already participated in this effort by submitting data for over 15 000 human proteins. The submitted data includes mass spectrometry and protein microarray-derived data, among other data types. Finally, HPRD is also linked to a compendium of human signaling pathways developed by our group, NetPath (http://www.netpath.org/), which currently contains annotations for several cancer and immune signaling pathways. Since the last update, more than 5500 new protein sequences have been added, making HPRD a comprehensive resource for studying the human proteome.

The availability of human genome sequence has transformed biomedical research over the past decade. However, an equivalent map for the human proteome with direct measurements of proteins and peptides does not exist yet. Here we present a draft map of the human proteome using high-resolution Fourier-transform mass spectrometry. In-depth proteomic profiling of 30 histologically normal human samples, including 17 adult tissues, 7 fetal tissues and 6 purified primary haematopoietic cells, resulted in identification of proteins encoded by 17,294 genes accounting for approximately 84% of the total annotated protein-coding genes in humans. A unique and comprehensive strategy for proteogenomic analysis enabled us to discover a number of novel protein-coding regions, which includes translated pseudogenes, non-coding RNAs and upstream open reading frames. This large human proteome catalogue (available as an interactive web-based resource at http://www.humanproteomemap.org) will complement available human genome and transcriptome data to accelerate biomedical research in health and disease.

We have developed NetPath as a resource of curated human signaling pathways. As an initial step, NetPath provides detailed maps of a number of immune signaling pathways, which include approximately 1,600 reactions annotated from the literature and more than 2,800 instances of transcriptionally regulated genes - all linked to over 5,500 published articles. We anticipate NetPath to become a consolidated resource for human signaling pathways that should enable systems biology approaches.

Signaling pathways are orchestrated by post-translational modifications (PTMs) such as phosphorylation. However, pathway analysis of PTM datasets generated by mass spectrometry (MS)-based proteomics is typically performed at a gene-centric level due to the lack of appropriately curated PTM signature databases and bioinformatic tools that leverage PTM site-specific information. Here we present the first version of PTMsigDB, a database of modification site-specific signatures of perturbations, kinase activities and signaling pathways curated from more than 2,500 publications. We adapted the widely used single sample Gene Set Enrichment Analysis approach to utilize PTMsigDB, enabling PTM Signature Enrichment Analysis (PTM-SEA) of quantitative MS data. We used a well-characterized dataset of epidermal growth factor (EGF)-perturbed cancer cells to evaluate our approach and demonstrated better representation of signaling events compared to gene-centric methods. We then applied PTM-SEA to analyze the phosphoproteomes of cancer cells treated with cell-cycle inhibitors and detected mechanism-of-action specific signatures of cell cycle kinases. We also applied our methods to analyze the phosphoproteomes of PI3K-inhibited human breast cancer cells and detected signatures of compounds inhibiting PI3K as well as targets downstream of PI3K (AKT, MAPK/ERK) covering a substantial fraction of the PI3K pathway. PTMsigDB and PTM-SEA can be freely accessed at https://github.com/broadinstitute/ssGSEA2.0.

Plasma Proteome Database (PPD; http://www.plasmaproteomedatabase.org/) was initially described in the year 2005 as a part of Human Proteome Organization’s (HUPO’s) pilot initiative on Human Plasma Proteome Project. Since then, improvements in proteomic technologies and increased throughput have led to identification of a large number of novel plasma proteins. To keep up with this increase in data, we have significantly enriched the proteomic information in PPD. This database currently contains information on 10 546 proteins detected in serum/plasma of which 3784 have been reported in two or more studies. The latest version of the database also incorporates mass spectrometry-derived data including experimentally verified proteotypic peptides used for multiple reaction monitoring assays. Other novel features include published plasma/serum concentrations for 1278 proteins along with a separate category of plasma-derived extracellular vesicle proteins. As plasma proteins have become a major thrust in the field of biomarkers, we have enabled a batch-based query designated Plasma Proteome Explorer, which will permit the users in screening a list of proteins or peptides against known plasma proteins to assess novelty of their data set. We believe that PPD will facilitate both clinical and basic research by serving as a comprehensive reference of plasma proteins in humans and accelerate biomarker discovery and translation efforts.