Building a Foundation for the Human Proteome: The Role of the Human Proteome Organization Sam Hanash*,† University of Michigan, Department of Pediatrics, 1150 W. Medical Center Dr. MSRB-1, A520, Ann Arbor, Michigan 48109-0656 Received December 11, 2003
Proteomics holds both substantial promise and substantial challenges. For proteomics to bear fruit on a large scale from a disease investigation point of view, it is essential to build a solid foundation for the field. Given the magnitude of the challenges, it is necessary to build a foundation by bringing together the private and public sectors. The Human Proteome organization is promoting the field of proteomics by engaging in such an effort and is developing several major initiatives. Keywords: proteomics • informatics • liver • plasma • brain • antibody
Introduction The completion of the human genome and the development of bio-mass spectrometry, have had a substantial impact on the field of proteomics by providing a sequence based framework and a needed instrumentation for deciphering the human proteome in health and in disease. Proteomics is particularly suited for disease investigations because its applicability to cells, tissues, and biological fluids alike. Nevertheless, the task of uncovering clinically useful findings based on proteomic profiling represents a substantial challenge due to the vast range of protein abundance and the compartmentalization of proteins. Continued improvements in proteomics tools promise to accelerate the pace of discovery of biomarkers and of other proteins of clinical utility. Although the proteome is encoded for in the human genome, numerous aspects of proteins, from their structure and function to their disease related alterations, cannot simply be inferred from genomic analysis. As a result, it has become appealing to entertain concepts for a global project related to the human proteome that may be analogous to the genome project. A major objective of such a project would be to characterize all proteins encoded for in the human genome and determine their range of variation in health and in disease. However, from an operational point of view, there are vast differences between a project to sequence the human genome and a project to decipher the human proteome, because of numerous added layers of complexity resulting from the highly dynamic nature of the proteome, in contrast to the relatively static nature of the human genome. The human proteome is near infinite, given the myriad of human cell and tissue types, each with its own unique repertoire of proteins. Thus, it is enormously difficult to conceive of a project to determine for all human cell types, their protein constituents and their range of expression and post-translational modifications, their interactions with other proteins and their occurrence as part of complexes, †
HUPO president.
10.1021/pr034126h CCC: $27.50
2004 American Chemical Society
their sub-cellular localization, and most importantly their structure and their function, all of which being context dependent and relevant to our understanding of disease related alterations. The Need for an Organized Effort in Proteomics. Uncovering the wide range of protein alterations in disease states holds much promise for disease prevention, disease diagnostics, and therapeutics. To that end, we need to develop a knowledge base of the normal human proteome. Given the enormous complexity of the human proteome, it stands to reason that no individual institution, be it public or private, would have the needed resources to tackle the human proteome single handedly. The Human Proteome Organization (http:// www.HUPO.org) came into existence to help define achievable objectives to further our understanding of the human proteome (Figure). HUPO’s stated mission is to consolidate national and regional proteome organizations into a worldwide organization; to engage in scientific and educational activities to encourage the spread of proteomics technologies and to disseminate knowledge pertaining to the human proteome and that of model organisms. To help define and prioritize objectives, HUPO has organized numerous meetings in various countries, with participation by government and industry representatives as well as academicians. As a result, a number of international initiatives have been formulated with goals to develop resources for proteomics on one hand and to engage in projects of a biological nature on the other. The current status of HUPO initiatives is found on the HUPO.org website and is reviewed here. The Development of Resources for Proteomics. Although mass spectrometry has provided a “shot in the arm” for proteomics, numerous additional resources, and technologies are needed to facilitate the mining of the human proteome. There is a substantial need for informatics resources in proteomics for practically every aspect of the field. For example, the current approaches to the analysis of protein data are highly informal and nonstandardized. An important informaticsJournal of Proteome Research 2004, 3, 197-199
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reviews related effort that HUPO is involved in is aimed at developing and adopting standardized approaches that facilitate the across lab analysis of proteomics data. The HUPO Proteomics Standards Initiative (PSI) was launched at a HUPO meeting in Washington, DC in April 2002, with the aim of defining community standards for data representation in proteomics to facilitate data comparison, exchange, and validation (http://psidev.sourcefourge.net). The initial focus has been on protein-protein interaction data and mass spectrometry data. A number of studies have been undertaken to define protein interaction networks by different groups. However, integration of date from different groups is hampered by the fragmentation of the data and its deposition in multiple databases and in different formats. Therefore, the HUPO PSI has proposed a community standard data model for the representation and exchange of protein interaction data. These data model is supported by major protein interaction data providers. Progress has also been made in the development of common standards for data exchange in the field of mass spectrometry. HUPO informatics efforts will also extend to other aspects of proteomics such as quantitative protein expression analysis. They will also address the needs of the targeted tissue proteomics initiatives with respect to data collection, storage and dissemination. Another major resource related initiative is the development of genome scale protein capture agents. The plans under development consist of initially making antibodies for the proteins identified as part of the HUPO proteome projects described below and eventually making antibodies to all the proteins encoded for in the human genome. These antibodies will allow proteome scale studies to be done as in the case of antibody microarrays that assay proteins in a tissue or biological fluid. Two major execution sites for antibody production will be located in Europe and in China and additional facilities are currently being explored. A piloting phase for antibody production in China has already begun. HUPO Proteome Projects. Much effort has been devoted to identifying a limited number of tissues that would be the focus of projects to characterize in depth their protein constituents. The comprehensive proteomic characterization of these tissues and their component cell populations represents an ambitious undertaking that is intended to push the limits of proteomics technologies, through a cooperative international effort. These targeted proteome projects will be executed in phases commensurate with the capabilities of available technologies. Tissue proteome initiatives are currently focused around three proteome projects consisting of the Plasma Proteome Project (PPP), the Liver Proteome Project (LPP) and the Brain Proteome Project (BPP). Each of these initiatives has been the focus of several workshops to date. For example in the past year, the LPP has been the subject of meetings in Beijing, Bethesda and Montreal to formalize the objectives of the project and identify participating countries and laboratories. The first of the tissue targeted projects to be implemented is the PPP which is currently in its piloting phase. The scientific objectives of the PPP are outlined in Table 1. The pilot phase for the PPP has begun using standardized samples distributed to some 50 laboratories throughout the world and is expected to last approximately one year. Funds have been raised by HUPO for the coordination of the piloting phase, with equal contributions from industry and from the National Institutes of Health in the United States. The objec198
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Hanash Table 1. Scientific Objectives of the PPP 1. Comprehensive analysis of plasma protein constituents in normal humans in large cohorts of subjects. 2. Determination of the extent of variation in plasma proteins: -within populations in various countries -across various populations from around the world 3. Identification of biological sources of variation within individuals over time and assessment of the effect of: -age, sex, diet and lifestyle -common medications and common diseases Table 2. Aims of the Piloting Phase of the PPP 1. Compare a broad range of technology platforms for the characterization of proteins in human plasma and serum. Assess resolution, sensitivity, time, cost, and volumes of sample required. 2. Clarify the influence of various technical variables in specimen collection, handling, and storage. 3. Assess the need and feasibility of depleting the most abundant plasma proteins and the need for additives for protein stability. 4. Develop a database structure and repository for HUPO PPP results. 5. Establish international collaborations for later-phase characterization of the normal human plasma proteome in different ethnic groups. 6. Lay the groundwork through evaluation of technology platforms and specimen handling for future studies of circulating proteins(biomarkers) in health and disease. Table 3. Aims of the LPP 1. Collection and banking of human liver tissue specimens. 2. Characterization of the protein expression profile of human liver. 3. Elucidation of post-translational modifications of liver proteins. 4. Construction of protein interaction maps of human liver. 5. Localization of human liver proteins in cellular compartments. 6. Development of an antibody bank for human liver proteins. 7. Development of an ORF bank for human liver proteins. 8. Studies of liver disease with a focus on hepatitis, liver cancer and related pathologies.
tives of the piloting phase are outlined in Table 2. HUPO is planning to organize a Jamboree at the completion of the piloting phase to review the findings resulting from the work undertaken by participating laboratories and which is expected to result in a database of identified proteins in serum and plasma. Once the piloting phase is completed, selected technologies will be utilized using standardized protocols, to achieve the scientific objectives of the project. Although the characterization of plasma proteins is challenging because of the vast dynamic range of protein abundance, the plasma is far less complex than an organ such as the liver or brain. In parallel with the development of PPP, HUPO has initiated two organ specific proteome projects, namely the LPP and the BPP. The scientific objectives of the LPP are outlined in Table 3. The objectives of the pilot phase for the LPP are currently being formulated. The importance of the LPP is demonstrated by the decision of the government of China to become a major participant in the project with the allocation of some $30 M to the pilot phase of the LPP and a projected contribution of some $200 M for the execution phase. Other countries with important contributions to the project include France, Canada, and the United States. The BPP has been the subject of several workshops to define scientific objectives and to develop aims for a pilot phase, along
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Role of the Human Proteome Organization Table 4. Summary of Activities to Date Related to the BPP November, 2002 December, 2002
April, 2003 September, 2003 October, 2004
January, 2004 April, 2004 October 2004
Discussions around an BPP at the 1st HUPO World Congress in Versailles, France HUPO joint Brain Proteomics workshop with the US National Institute of Neurological Diseases and Stroke, Washington, DC BPP kick-off meeting in Frankfurt, Germany HUPO BPP Workshop, Germany Presentation of the HUPO BPP concept at the 2nd HUPO World Congress in Montreal, Canada, and appointment by HUPO of a planning committee for BPP. Meeting of the BPP Planning Committee in Paris, France HUPO BPP Workshop, Paris, France Symposium and Workshop at the 3rd HUPO World Congress in Beijing, launching BPP
similar lines as the LPP. A planning committee has been constituted to formally organize the project, with Germany playing a major role at the present time. Table 4 lists the activities undertaken to date to lay a foundation for the BPP. The Need for Education and Training and Scholarly Activities in Proteomics. In addition to regional meetings and workshops, annual gatherings to review the progress in the field that attract established investigators, postdoctoral fellows and graduate students have been organized. HUPO’s first annual proteomics congress which took place in Versailles, France in November, 2002 was highly successful and attracted some 1000 participants, HUPO’s equally highly successful second annual congress which took place in Montreal attracted some 2500 participants. These annual congresses are scheduled to take place on a rotating basis in different continents, with the next HUPO annual congress being scheduled for Beijing, China, October 25-28, 2004. The Beijing congress will include in addition to a scientific and edudcational program, separate workshops devoted to each of the different HUPO initiatives.
Figure 1. Organization of HUPO into three major spheres: HUPO Americas, HUPO Europe and HUPO Asia/Oceania. Countries currently represented in HUPO Asia/Oceania are shown.
Scholarships will be available to encourage participation by young investigators. HUPO will also engage in an extensive educational and training effort in proteomics. Plans to this effect are currently being sketched out. HUPO will organize educational seminars, courses and training workshops throughout the world to educate a new generation of scientists in this emerging field. HUPO will also jointly sponsor similar activities with other public or private organizations. It is envisioned that a web based proteomics portal will be developed that makes accessible easy read overviews of various aspects of proteomics as well as reviews of recent developments and relevant technologies. Programs from short intensive courses to extended laboratory visits will be organized. PR034126H
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