Src tyrosine kinase-induced loss of luteinizing hormone responsiveness is via a Ras-dependent, phosphatidylinositol-3-kinase independent pathway.
Gonadotropins stimulate gonadal cell steroid secretion primarily through activation of a cAMP-protein kinase A signal transduction pathway. Various growth factors have been shown to inhibit gonadotropin-stimulated steroidogenesis, however, the intracellular signaling cascades involved in growth factor inhibition have not been characterized. The present study investigated whether Src tyrosine kinase, a nonreceptor tyrosine kinase activated in response to growth factor stimulation and previously shown to inhibit LH-stimulated progesterone secretion, acts via activation of Ras stimulated pathways, phosphatidylinositol-3-kinase (PI3-kinase) stimulated pathways, or both in MA10 Leydig cells. Direct activation of Src in MA10 cells that express a temperature sensitive Src was associated with an increase in GTP-bound Ras, indicating increased Ras activity in response to Src activation. Direct activation of Ras by way of expression of a constitutively active Ras (Ras+) was associated with a decrease in LH responsiveness. Coexpression of a dominant negative Src, which by itself increases LH responsiveness in MA10 cells, had no effect on Ras+ inhibition on LH responsiveness, further demonstrating that Src is upstream of Ras. In addition, MA10(Ras+) cells were relatively unresponsive to cholera toxin or 8-bromo cAMP, indicating the effects of Ras are independent of cAMP generation. Wortmannin, a PI3-kinase inhibitor, did not restore LH responsiveness to cells expressing activated Src or constitutively active Ras. These results demonstrate that Src activates a Ras pathway in MA10 Leydig cells, and that activation of Ras is associated with a loss of LH responsiveness that is independent of PI3-kinase.
Taylor
Christopher C
CC
Department of Cell Biology, Georgetown University School of Medicine, Washington, District of Columbia 20007, USA. cct5@georgetown.edu
D000730:Androstadienes / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D006153:Guanosine Diphosphate / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000378:metabolism; D007984:Leydig Cell Tumor; D007986:Luteinizing Hormone / Q000494:pharmacology*; D016296:Mutagenesis; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011374:Progesterone / Q000378:metabolism; D015398:Signal Transduction; D014162:Transfection; D014407:Tumor Cells, Cultured; D000077191:Wortmannin; D018631:ras Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D019061:src-Family Kinases / Q000235:genetics / Q000378:metabolism*
D000730:Androstadienes; D004791:Enzyme Inhibitors; D006153:Guanosine Diphosphate; D006160:Guanosine Triphosphate; D007986:Luteinizing Hormone; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011374:Progesterone; D000077191:Wortmannin; D018631:ras Proteins; D019061:src-Family Kinases