The human proteome project: current state and future direction

After the successful completion of the Human Genome Project, the Human Proteome Organization has recently officially launched a global Human Proteome Project (HPP), which is designed to map the entire human protein set. Given the lack of protein-level evidence for about 30% of the estimated 20,300 protein-coding genes, a systematic global effort will be necessary to achieve this goal with respect to protein abundance, distribution, subcellular localization, interaction with other biomolecules, and functions at specific time points. As a general experimental strategy, HPP research groups will use the three working pillars for HPP: mass spectrometry, antibody capture, and bioinformatics tools and knowledge bases. The HPP participants will take advantage of the output and cross-analyses from the ongoing Human Proteome Organization initiatives and a chromosome-centric protein mapping strategy, termed C-HPP, with which many national teams are currently engaged. In addition, numerous biologically driven and disease-oriented projects will be stimulated and facilitated by the HPP. Timely planning with proper governance of HPP will deliver a protein parts list, reagents, and tools for protein studies and analyses, and a stronger basis for personalized medicine. The Human Proteome Organization urges each national research funding agency and the scientific community at large to identify their preferred pathways to participate in aspects of this highly promising project in a HPP consortium of funders and investigators.     

Human liver proteome project: plan, progress, and perspectives

The Human Liver Proteome Project is the first initiative of the human proteome project for human organs/tissues and aims at writing a modern Prometheus myth. Its global scientific objectives are to reveal the "solar system" of the human liver proteome, expression profiles, modification profiles, a protein linkage (protein-protein interaction) map, and a proteome localization map, and to define an ORFeome, physiome, and pathome. Since it was first proposed in April 2002, the Human Liver Proteome Project has attracted more than 100 laboratories from all over the world. In the ensuing 3 years, we set up a management infrastructure, identified reference laboratories, confirmed standard operating procedures, initiated international research collaborations, and finally achieved the first set of expression profile data.     

"Does understanding the brain need proteomics and does understanding proteomics need brains?"--Second HUPO HBPP Workshop hosted in Paris

The second Human Brain Proteome Project (HBPP) Workshop of the Human Proteome Organisation (HUPO) took place at the Ecole Supérieure de Physique et de Chimie Industrielles de la Ville de Paris (ESPCI) from April 23-24, 2004. During two days, more than 70 attendees from Europe, Asia and the US came together to decide basic strategic approaches, standards and the beginning of a pilot phase prior to further studies of the human brain proteome. The international consortium presented the technological and scientific portfolio and scheduled the time table for the next year.     

The human eye proteome project

The human eye proteome is the latest addition to the HUPO Human Proteome Project (HPP). Semba et al. (The Human Eye Proteome Project: Perspectives on an emerging proteome. Proteomics 2013, 13, 2500–2511) establish a provisional baseline for the proteomes of the many anatomical compartments of the eye, based on literature review. As part of the Biology and Disease‐driven HPP, they and their colleagues will generate fresh data and meet the stringent guidelines for protein identification and characterization as established by the HPP.     

Towards standard protocols and guidelines for urine proteomics: a report on the Human Kidney and Urine Proteome Project (HKUPP) symposium and workshop, 6 October 2007, Seoul, Korea and 1 November 2007, San Francisco, CA, USA

The Human Kidney and Urine Proteome Project (HKUPP) was initiated in 2005 to promote proteomics research in the nephrology field, to better understand kidney functions as well as pathogenic mechanisms of kidney diseases, and to define novel biomarkers and therapeutic targets. This project was first approved in 2005 by the Human Proteome Organisation (HUPO) as a Kidney Disease Initiative under an umbrella of the HUPO Disease Biomarker Initiative (DBI), and more recently was approved as the HKUPP Initiative in 2007. Several sub-projects have been planned to achieve the ultimate goals. The most pressing is the establishment of "standard protocols and guidelines for urine proteome analysis". This sub-project had been extensively discussed during the first HKUPP symposium (during 6(th) HUPO Annual World Congress--October 2007, Seoul, Korea) and second workshop (during 40(th) American Society of Nephrology Renal Week--November 2007, San Francisco, CA, USA). Additional data and references have been collected after the symposium and workshop. An initial draft of standard protocols and guidelines for proteome analysis of non-proteinuric urine (urine protein excretion < or =150 mg/day) will soon be released as the first HKUPP product.     

Automated reprocessing pipeline for searching heterogeneous mass spectrometric data of the HUPO Brain Proteome Project pilot phase

The newly available techniques for sensitive proteome analysis and the resulting amount of data require a new bioinformatics focus on automatic methods for spectrum reprocessing and peptide/protein validation. Manual validation of results in such studies is not feasible and objective enough for quality relevant interpretation. The necessity for tools enabling an automatic quality control is, therefore, important to produce reliable and comparable data in such big consortia as the Human Proteome Organization Brain Proteome Project. Standards and well-defined processing pipelines are important for these consortia. We show a way for choosing the right database model, through collecting data, processing these with a decoy database and end up with a quality controlled protein list merged from several search engines, including a known false-positive rate.     

Chromosome-centric approach to overcoming bottlenecks in the Human Proteome Project

The international Human Proteome Project (HPP), a logical continuation of the Human Genome Project, was launched on 23 September 2010 in Sydney, Australia. In accordance with the gene-centric approach, the goals of the HPP are to prepare an inventory of all human proteins and decipher the network of cellular protein interactions. The greater complexity of the proteome in comparison to the genome gives rise to three bottlenecks in the implementation of the HPP. The main bottleneck is the insufficient sensitivity of proteomic technologies, hampering the detection of proteins with low- and ultra-low copy numbers. The second bottleneck is related to poor reproducibility of proteomic methods and the lack of a so-called ‘gold’ standard. The last bottleneck is the dynamic nature of the proteome: its instability over time. The authors here discuss approaches to overcome these bottlenecks in order to improve the success of the HPP.     

血液蛋白质组学的研究技术及进展

人血液含有来源于几乎所有细胞、组织、器官的蛋白质,可以直接反映病理、生理状态,是各种疾病诊断、生物标志物发现的最有价值的标本。因此,长期以来,血浆蛋白质组一直是人们研究的热点,并被人类蛋白质组组织(HUPO)列为首批启动的重大国际合作研究项目。血浆蛋白质含量动态范围非常广、成分极其复杂,血浆蛋白质组的研究极富挑战性。近年来,血浆高丰度蛋白质去除、蛋白质/肽段分离、质谱鉴定、数据处理等多种相关技术都取得了很大的进展。本文简要综述了上述技术领域的研究和应用进展。     

Ian Humphery-Smith on current challenges in proteomics

Ian Humphery-Smith is Professor of Pharmaceutical Proteomics at Utrecht University, The Netherlands, and until recently was a Managing Director and Chief Scientific Officer of Glaucus Proteomics. After a PhD in Parasitology at the University of Queensland, he studied virology and bacteriology in France as a post-doc, before returning to Australia as Course-Coordinator in Medical Microbiology and Immunology at the University of Sydney. During this time, Humphery-Smith took up the posts of Executive Director of Australia's second largest DNA sequencing facility and Director of the Center for Proteomic Research and Gene-Product Mapping, which later became the world's first center to focus on studying the proteome. Humphery-Smith has devoted ten years of research to analyzing proteins in health and disease, and it was his work that originally coined the term ‘proteomics’. He was the first to publish the most complete analysis of an entire proteome in 2000, that of the bacterium Mycoplasma genitalium. He currently serves as a council member of the Human Proteome Organization (HUPO) and has been a prime mover in efforts to have the Human Proteome Project become a formally-ratified international initiative to follow-on from the Human Genome Project.     

A comparison of the HUPO Brain Proteome Project pilot with other proteomics studies

The pilot phase of the Brain Proteome Project (BPP), the Human Proteome Organization (HUPO) initiative that focuses on studies of the brain of both humans and mice, has now been completed. Participating laboratories studied the proteomes of two human samples derived from biopsy and autopsy as well as three mouse samples from various developmental stages. With the combined and centrally reprocessed data now available, a comparison in terms of protein identifications and project organization is made between the HUPO BPP pilot and three other proteomics studies: the HUPO Plasma Proteome Project (PPP) pilot, a proteome of human blood platelets and a recently published comprehensive mouse proteome. Finally, as any comparison between large-scale proteomics datasets is decidedly non-trivial, we also evaluate and discuss several ways to go about comparing such different result sets.     

The HUPO Plasma Proteome Project: a report from the Munich congress

The Human Proteome Organization has several major collaborative research initiatives, including the Plasma Proteome Project. A major feature of the HUPO World Congress in Munich in August 2005 was the release of the special issue of PROTEOMICS with 28 articles from the pilot phase of the Plasma Proteome Project. An open Workshop and a presentation in the closing plenary session of the congress focused on next phases for the Plasma Proteome Project.     

Report of the 9th HLPP Workshop October 2007, Seoul, Korea

The Human Liver Proteome Project is one of the Human Proteome Initiatives launched by Human Proteome Organization (HUPO). Major achievements of the project have been obtained under the efforts of international collaboration with all the participants since it was formally proposed in 2002. Its updated progresses were presented in the latest workshop held in conjunction with the sixth HUPO World Congress in October, 2007, Seoul, Korea. Furthermore, four topics related to the project as well as other initiatives were lively discussed among all the attendees.     

Proteomic analysis of individual variation in normal livers of human beings using difference gel electrophoresis

Normal Chinese Liver Proteome Expression Profile is one of the major parts of Human Liver Proteome Project. Before starting the studies, it is necessary to examine the interindividual variation of normal liver proteome and evaluate the minimal size of samples for proteomic analysis. In this study, normal liver samples from ten individual volunteers were collected and the proteome profiles of these samples were analyzed using 2-D difference gel electrophoresis (DIGE) combined with MALDI-TOF/TOF MS. The individual liver tissue lysates were labeled with Cy3 and Cy5 while the pooled sample was labeled with Cy2 as an internal standard, which minimized gel-to-gel variation. After analysis by the DeCyder software, up to 2056 protein spots were detected on the master gel. The CV of standardized abundance was calculated for the protein spots that were matched across all ten gels. The CV values of these protein spots ranged from 6.4 to 108.5% and the median CV was approximately 19%, which demonstrated that the protein expression of normal liver among different individuals was relatively stable. The eight proteins with CV values over 50% were identified which would be a caveat when considering these proteins as potential disease-related markers. Moreover, the one-way ANOVA feature showed a correlation between sample size and individual variations. The results showed that when the sample size exceeded 7, the individual variations were not significant to the whole pool. Our results are an important basis for liver protein expression profiles and comparative proteomics of liver disease.     

Age-related proteome analysis of the mouse brain: a 2-DE study

2-DE remains the most popular and versatile protein separation method among a rapidly growing array of various proteomics technologies. However, variability in sample processing, experimental design and data analyses results in a limited cross-validation between studies performed in different laboratories. One of the goals of the Human Proteome Organization (HUPO) is to establish standards and guidelines for proteomics studies. We contributed to the HUPO Brain Proteome Project by analyzing brains from neonatal and adult mice using 2-DE. Here we propose a standard workflow to analyze 2-DE images and extract statistically significant differences. After differential analysis and identification by MALDI-TOF/TOF, dihydropyrimidinase-related proteins, brain FABP, stathmin, isocitrate dehydrogenase, gamma enolase, annexin V, glutamine synthetase, creatine kinase B chain, triosephosphate dehydrogenase, and malate dehydrogenase were found differentially expressed between the two groups. The functions and potential mechanisms underlying the variation observed for these proteins are discussed.     

Analysis of the HUPO Brain Proteome reference samples using 2-D DIGE and 2-D LC-MS/MS

Within the Human Proteome Organization (HUPO) Brain Proteome Project, a pilot study was launched with reference samples shipped to nine international laboratories (see Hamacher et al., this Special Issue) to evaluate different proteome approaches in neuroscience and to build up a first version of a brain protein database. One part of the study addresses quantitative proteome alterations between three developmental stages (embryonic day 16; postnatal day 7; 8 weeks) of mouse brains. Five brains per stage were differentially analyzed by 2-D DIGE using internal standardization and overlapping pH gradients (pH 4-7 and 6-9). In total, 214 protein spots showing stage-dependent intensity alterations (> two-fold) were detected, 56 of which were identified. Several of them, e.g. members of the dihydropyrimidinase family, are known to be associated with brain development. To feed the HUPO BPP brain protein database, a robust 2-D LC-MS/MS method was applied to murine postnatal day 7 and human post-mortem brain samples. Using MASCOT and the IPI database, 350 human and 481 mouse proteins could be identified by at least two different peptides. The data are accessible through the PRIDE database (http://www.ebi.ac.uk/pride/).     

Evaluation of 2-DE protein patterns from pre- and postnatal stages of the mouse brain

Brains of the mouse from three developmental stages, embryo day 16 (Ed16), postnatal stage one week (1W) and eight weeks (8W), were distributed to different laboratories for a collaborative proteome analysis (The Human Brain Proteome Project). As one of the laboratories involved in this project, we separated total protein extracts of the brains by large gel 2-DE. From the 2-DE protein patterns a section was evaluated for each of the three stages according to resolution, reproducibility and quantitative changes using an image analysis software. The evaluated pattern section was selected to allow comparisons of 2-DE patterns between different laboratories on the basis of optimum separation. Changes in protein expression were analysed within two phases of development: Stage Ed16 versus stage 1W and stage 1W versus stage 8W. Out of the 200 protein spots evaluated 5-6% showed quantitative changes in the range of > or = 30% between two stages. The relationship in the frequency of up- and down-regulated protein spots differed between the two investigated phases. Most of the protein spots which showed altered expression between two stages were identified by MS. High quality in protein separation and evaluation is demonstrated.     

HUPO Brain Proteome Project: summary of the pilot phase and introduction of a comprehensive data reprocessing strategy

The Human Proteome Organisation (HUPO) initiated several projects focusing on the proteome analysis of distinct human organs. The Brain Proteome Project (BPP) is the initiative dedicated to the brain, its development and correlated diseases. Two pilot studies have been performed aiming at the comparison of techniques, laboratories and approaches. With the help of the results gained, objective data submission, storage and reprocessing workflow have been established. The biological relevance of the data will be drawn from the inter-laboratory comparisons as well as from the re-calculation of all data sets submitted by the different groups. In the following, results of the single groups as well as the centralised reprocessing effort will be summarised and compared, showing the added value of this concerted work.     

17th Chromosome‐Centric Human Proteome Project Symposium in Tehran

This report describes the 17th Chromosome‐Centric Human Proteome Project which was held in Tehran, Iran, April 27 and 28, 2017. A brief summary of the symposium's talks including new technical and computational approaches for the identification of novel proteins from non‐coding genomic regions, physicochemical and biological causes of missing proteins, and the close interactions between Chromosome‐ and Biology/Disease‐driven Human Proteome Project are presented. A synopsis of decisions made on the prospective programs to maintain collaborative works, share resources and information, and establishment of a newly organized working group, the task force for missing protein analysis are discussed.