Prolactin (PRL) receptors regulate vital physiological processes such as reproduction, immune function, morphogenesis, cell proliferation and differentiation. Human PRL receptors are activated by PRL as well as placental lactogen and growth hormone. Overstimulation of PRL receptors has been associated with breast and prostate tumor promotion. the long-term objective of this proposal is to understand the molecular basis for how prolactin receptors activate multiple parallel signal transduction pathways in target cells. As our first major objective in pursuit of this goal, we proposed on the basis of work that will presented under Preliminary Studies to test the central hypothesis that prolactin receptors signal via multiple parallel transduction pathways that are cell-dependent and selectively modulated by external stimuli. This proposal has three major, intimately linked goals, each maintaining a primary focus processes. These goals are: 1) To determine how PRL receptors interact with multiple JAK tyrosine kinases (JAK1, JAK2 and TYk2) in a cell dependent manner, 2) to identify PRL receptor-mediated activation of Src-tyrosine kinases in mammary epithelial cells and hematopoietic cells, and investigate the importance o cytoplasmic receptor regions for cell-dependent recruitment of specific Src kinases and the activation of the SHC/RAS/MAPK pathway, 3) to determine whether activation of multiple parallel signaling pathways by PRL in target cells can be selectively regulated by external physiological and pharmacological modulators, and specifically explore the impact of maintained JAK2/STAT5a/b signals and uncoupling of STAT1/3SHC- RAS-MAPK signals on PRL inducted proliferation and gene regulation in Nb2 lymphoctyes. The best models of direct studies of these initial biochemical reactions remain simple, homogenous cell populations cultured in vitro. Rat Nb2 pre-T lymphocytes and several well-differentiated human mammary epithelial cells lines will be used for these studies. PRL signaling will also be reconstituted in naive myeloid cells and b the introduction of novel PRL receptor variants into mammary epithelial cells. Our expectations are that, at the conclusion of the proposed period of support, we will have identified how PRL receptors can activate multiple JAKs, and defined regions of the human PRL receptor specifically responsible for triggering SHC/RAS/MAPK and Src tyrosine kinases in relevant human mammary cells. We also expect to shed new light on external modulation of individual PRL receptor signaling pathways. With our extensive background in signal transduction, we expect to converge to regulate vital physiological processes such as cellular growth and differentiation.
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