This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Recent technological developments enable two complementary approaches to mapping the signaling and regulatory circuitry inside cells. First, it is possible to directly measure the wires themselves, by screening for protein-protein and protein-DNA interactions using systematic protein (Gavin et al., 2002) and chromatin immunoprecipitation (Ren et al., 2000) experiments. Second, we can measure the molecular and cellular states induced by the wiring. For example, changes in gene expression are measured with DNA microarrays (DeRisi et al., 1997), while changes in protein abundance (Gygi et al., 1999), protein phosphorylation state (Zhou et al., 2001), and metabolite concentrations (Griffin et al., 2001) may be quantitated with mass spectrometry, NMR, and other advanced techniques. In this proposal we combine both of these approaches in a multi-tiered pathway mapping strategy. First, large databases of protein-protein and protein-DNA interactions are used to define a global network or wiring scaffold . Second, this scaffold is filtered against changes in mRNA expression, protein expression, and post-translational modifications (i.e., phosphorylation) recorded in response to different cellular perturbations. Regions of the wiring scaffold whose mRNA or protein states are significantly activated by perturbation are identified and mapped according to a computational search algorithm. The interaction pathways and complexes making up each activated region in the scaffold become prime candidates for further verification and modeling as important signaling and compensatory mechanisms controlling the cell s perturbation response.
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