The goal of the proposed studies is to understand how the structure of the Src tyrosine kinase regulates its function as a signaling molecule. Src is a member of a family of tyrosine protein kinases which play critical roles in both normal and malignant cell growth. The proposal has two specific aims.
Aim 1 will focus on signaling by members of the Src family using three lines of investigation. First, a novel Src SH3 binding protein identified by the PI during the previous grant period, termed Dif-1, will be examined for its role in adipose differentiation and intracellular signaling. Second, the role of tyrosine phosphorylation of Raf-1 will be probed. Third, a modified version of the yeast 2-hybrid system, in which a tyrosine kinase is also introduced, will be exploited in an attempt to identify new substrates for Src kinases.
Aim 2 will examine the structure of the Lck tyrosine kinase in great detail, using a synthetic lck gene generated by the PI. A specific region within the N-terminus of kinase subdomain VII will be targeted for mutagenesis, in an attempt to identify Lck mutants with altered substrate specificity. Mutants will be initially screened by identifying alterations in tyrosine phosphorylation patterns in E. coli, followed by peptide library screening. A second set of experiments will examine the function of the """"""""activation loop"""""""", a region downstream from the tyrosine autophosphorylation site of Lck. The PI will look for SH2 containing proteins which bind to this region.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Cellular Biology and Physiology Subcommittee 1 (CBY)
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Spalholz, Barbara A
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Dana-Farber Cancer Institute
United States
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Laham, L E; Mukhopadhyay, N; Roberts, T M (2000) The activation loop in Lck regulates oncogenic potential by inhibiting basal kinase activity and restricting substrate specificity. Oncogene 19:3961-70
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