Hyperprolactinemia is an important cause of hypogonadism and infertility in men and women. Prolactin (PRL) receptors have been identified in many tissues, but how PRL causes these clinical effects and others are not well understood because what events follow PRL receptor binding in most tissues are still controversial. This grant proposal aims to investigate some of the biochemical events that may follow PRL binding in a known target cell (the Nb2 lymphoma cell) which demonstrates a biologic response specific to lactogenic hormones (cell growth and division). Previously, it has been shown that bacterial toxins, pertussis toxin, (islet activating protein: IAP) and cholera toxin (CT), can modify PRL-stimulated mitogenesis in the Nb2 cell. These studies suggest that one or both toxins may mediate their predominantly inhibitory actions on the Nb2 cell through generation of increased cellular concentrations of cyclic 3'5'- adenosine monophosphate (cAMP). However, these toxins also affect cellular processes unrelated to the generation of cAMP, such as, for example, the ADP- ribosylation of non-adenylate cyclase associated GTP binding proteins (G proteins). Thus, it is also possible that either toxin, or both, alter PRL-stimulated mitogenesis in the Nb2 cell through one of these mechanisms. This proposal will test the specific hypothesis that either IAP, CT, or both toxins alter PRL- stimulated mitogenesis in the Nb2 cell because, through ADP ribosylation, they are able to alter the function of one or more G proteins involved in PRL signal transduction. In order to prove the importance of G proteins in PRL action, it will be necessary to demonstrate two or more of the following: 1) IAP and/or CT stimulate ADP ribosylation CT in Nb2 membranes; 2) GTP or its analogs alters binding of lactogenic hormone to its receptors; 3) PRL alters binding of /gtp or its analog to Nb2 membranes; 4) PRL enhances GTPase activity; or 5) activated GTP or its analog mimics the action of PRL in membrane or cells. This last possibility is not yet demonstrable as no second messenger or other biochemical activity specific to PRL action has yet been discovered. If the hypothesis is proven correct, the proposal would then lead to the isolation and characterization of the G protein(s) involved, including to what second messenger systems it may be linked.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
First Independent Research Support & Transition (FIRST) Awards (R29)
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Reproductive Endocrinology Study Section (REN)
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University of Nebraska Medical Center
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