The objectives of this proposal are threefold: a) to investigate-aspects of c-kit mediated signaling in hematopoietic stem cell populations, b) to investigate the respective role of the soluble and the cell membrane forms of the ligand KL of the c-kit receptor and c) to determine the significance of co-expression of the c-kit receptor and kit-ligand, KL i.e. autocrine mechanisms. C-kit is the cellular homolog of the oncogene v-kit and encodes a receptor tyrosine kinase which belongs to the platelet-derived growth factor (PDGF) receptor subfamily (Besmer et al., 1986). We and others have identified and characterized the ligand of the c-kit receptor we designated KL. The KL gene encodes a cell surface molecule which may be proteolytically processed to produce a soluble product similar or identical to the one we had purified. While the c-kit gene is allelic with the murine white spotting locus (W), the KL gene was shown to be allelic with the murine steel locus. Mutations at both the W and the Sl loci affect various aspects of hematopoiesis, including the stem cell hierarchy, and the proliferation and/or migration of primordial germ cells and melanoblasts during development. Based on the analysis of mutant phenotypes the cellular responses the c-kit receptor appears to mediate are quite diverse and include: cell proliferation, cell survival, cell migration, cell differentiation and other post mitotic functions. The elucidation of the mechanisms of these different responses is of fundamental importance. In part, the current proposal is designed to address issues concerning the mechanism of c*it mediated survival and proliferation (self renewal) in hematopoietic stem cell populations. The cell-membrane bound nature of KL poses important questions about its function and mode of action. The current proposal is designed to address some of these issues. The goals toward this end include: l). An analysis of KL processing and elucidation of the function of soluble and membrane bound KL. 2) The in vivo analysis of mutant versions of KL in transgenic and chimeric mice. 3) The construction of mice expressing KL-2 but not KL- 1 by germline modification. 4) The characterization of the hematopoietic stem cell compartment in Sl mutant mice. 5) The derivation of transgenic mice expressing human versions of KL to facilitate engraftment and development of human hematopoietic cells in mouse models. 6) The evaluation of autocrine mechanisms involving cell membrane associate and soluble forms of KL in in vitro cell culture systems and in in vivo models.

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Research Project (R01)
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Hematology Subcommittee 2 (HEM)
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Sloan-Kettering Institute for Cancer Research
New York
United States
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