The goal of this research project is to determine how proteins assemble into complexes that function to direct development and adaptive responses. The central focus is on LIM domains, Zn2+ binding protein modules located in both nuclear and cytoplasmic proteins, that act to assemble molecular complexes. LIM domain containing proteins are essential for nervous system, hematopoietic and muscle development; they are also essential for cytoskeletal function.
Specific aims i nclude: 1) Determine the structural basis of molecular recognition by LIM domains. Nuclear LIM interactor (NLI) is the major high affinity target recognized by LIM homeodomain (LH) and LIM only (LMO) proteins. Goals are to a) determine features of NLI involved in LIM domain binding, b) determine features of nuclear LIM domains necessary for high affinity molecular recognition, c) determine the kinetics and stoichiometry of NLI binding and d) determine the structure of a nuclear LIM domain NLI complex. 2. Analyze the function of nuclear LIM interactor. Predictions based on biochemical studies will be critically tested in functional assays. Goals are to a) analyze the function of NLI in retinoic acid-induced neuronal differentiation of P19 cells and b) analyze the function of NLI in transcription mediated by LH and LMO proteins. 3. Determine the function of nuclear LIM only proteins. These studies will focus on a newly discovered protein LM04. Goals are to a) determine the expression of LM04 during mouse development and compare it to LH, LMO and transcription factors essential for specific developmental processes. b) determine the effect of targeted disruption of LM04 in mouse development. 4. Analyze the function of cytoplasmic LIM domains. Many cytoplasmic LIM domain proteins regulate the cytoskeleton and have molecular recognition that is distinct from nuclear LIM domains. Goals are to a) identify and characterize high affinity interactions that involve cooperation between cytoplasmic LIM domains and b) determine the function of the LIM domains of LIM kinase. Regulation of LIM kinase that controls features of the actin cytoskeleton will be defined.
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