This revised competitive renewal of an R24 Resource-Related Research Grant is focused on creating proteomic resources that can be applied to research using nonhuman primates. The proposal builds upon resources constructed during the original funding period in which we developed extensive rhesus macaque (Macaca mulatta)-specific genomic tools. Through the construction of complementary proteomic resources, we intend to provide the research community with the full spectrum of species-specific genomic and proteomic tools and databases needed to apply the technologies of functional genomics to nonhuman primate research. To build these resources, we have teamed with the Biological Systems Analysis and Mass Spectrometry group headed by Dr. Richard D. Smith at Pacific Northwest National Laboratory. This group brings to the effort unrivaled mass spectrometry instrumentation and cutting-edge proteomics expertise.
In Specific Aim 1, we will use shotgun proteomics and Fourier transform ion cyclotron resonance mass spectrometry to build macaque specific protein databases for use in identifying proteins present in various rhesus macaque tissues. These databases will be used in Aim 2 to characterize the changes in protein profiles that occur in response to simian immunodeficiency virus (SIV) infection. We will begin by analyzing peripheral blood mononuclear cells (PBMCs) infected in vitro with SIVmac239. We will then infect the same animals from which the PBMCs were derived and analyze blood, gut-associated lymphoid tissue, and lymph nodes at multiple early time points after infection. Quantitative proteomic analyses will be performed using stable isotope labeling and a label-free comparative absolute quantification method in combination with the AMT tag approach. These studies will serve to drive the development of proteomic resources and to enhance our understanding of the use of macaques as models for AIDS research.
In Aim 3, we will develop the bioinformatics infrastructure needed for the storage, interpretation, and dissemination of proteomics data, incorporating many open source tools including LabKey. This information resource will offer Web access to tiers of results throughout the process. As part of our efforts to extend the use of proteomics to the nonhuman primate research community, we will also host annual workshops where investigators can discuss how to make use of proteomic databases and to incorporate proteomics into their own research activities.
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