Peter Besmer: Mechanism of Kit Signaling

Role of Kit Signaling through PI 3-kinase and SRC Kinase Pathways in Mast Cells

The receptor tyrosine kinase Kit plays critical roles in hematopoiesis, gametogenesis, and melanogenesis. In mast cells, Kit receptor activation mediates several cellular responses, including cell proliferation and suppression of apoptosis induced by growth factor deprivation and gamma-irradiation. Kit receptor functions are mediated by kinase activation, receptor autophosphorylation, and association with various signaling molecules.

We have investigated the role of PI 3-kinase and Src kinases in Kit-mediated cell proliferation and suppression of apoptosis induced both by factor deprivation and irradiation in bone marrow-derived mast cells (BMMC). Analysis of Kit-/- BMMC expressing mutant Kit receptors and the use of pharmacological inhibitors revealed that both signaling pathways contribute to these Kit-mediated responses and that elimination of both pathways abolishes them.

We demonstrated that the PI 3-kinase and Src kinase signaling pathways converge to activate Rac1 and JNK. Analysis of BMMC expressing wild type and dominant-negative mutant forms of Rac1 and JNK revealed that the Rac1/JNK pathway is critical for KL-induced proliferation of mast cells but not for suppression of apoptosis. In addition, KL was shown to inhibit sustained activation of JNK induced by gamma-irradiation and concomitant irradiation-induced apoptosis.

Point Mutation in Kit Receptor Tyrosine Kinase Reveals Essential Roles for Kit Signaling in Oogenesis and Spermatogenesis without Affecting Other Kit Responses

In mast cells, Kit-mediated recruitment and activation of PI 3-kinase produces phosphatidylinositol 3-phosphates and plays a critical role in mediating cell adhesion and secretion, and has contributory roles in mediating cell survival and proliferation. To investigate the consequences in vivo of blocking Kit-mediated PI 3-kinase activation, we have mutated the binding site for the p85 subunit of PI 3-kinase in the c-kit gene, using a knock-in strategy. Mutant mice have no pigment deficiency or impairment of steady state hematopoiesis. However, gametogenesis is affected in several ways, and tissue mast cell numbers are affected differentially. While primordial germ cells during embryonic development are not affected, KitY719F/KitY719F males are sterile due to a block at the premeiotic stages in spermatogenesis. Furthermore, adult males develop Leydig cell hyperplasia. The Leydig cell hyperplasia implies a role for Kit in Leydig cell differentiation and/or steroidogenesis.

In mutant females, follicle development is impaired at the cuboidal stages resulting in reduced fertility. Also, adult mutant females develop ovarian cysts and ovarian tubular hyperplasia. Therefore, a block in Kit receptor mediated PI 3-kinase signaling may be compensated for in hematopoiesis, melanogenesis, and primordial germ cell development, but is critical in spermatogenesis and oogenesis.