The role and regulation of cell survival in uterine functions.

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BACKGROUND & RATIONALE: Prostate apoptosis response-4 (Par-4), a novel regulator of apoptosis as well as epithelial to mesenchymal transition (EMT), is active in a variety of cell lines and knockout mice show reduced lifespan and enhanced abnormal cell proliferation in the endometrium. We demonstrated that Par-4 is a direct target of caspase-3 during apoptosis induction and that the resulting cleaved fragment is capable of inducing programmed cell death through its translocation to nucleus. We also showed that TGFß increases EMT of endometrial cells and that it can regulate the long form of Par-4 in vitro. Interestingly, preliminary evidence gathered using cell lines in our laboratory and publically available databases suggest that 17ß-estradiol (E2) can negatively regulate Par-4. E2, an important female sex hormone, act through two known pathways, the canonical genomic pathway involving its intracellular receptor ER? which bind to estrogen-response elements (ERE) in DNA and the non-genomic pathway involving the PI-3K/Akt signaling action, an important survival and proliferation actor in the endometrium. How E2 action can be modulated through the inhibition of the pro-apoptotic factor Par-4 is unknown and it is crucial that we understand the molecular mechanisms controlling these functions. HYPOTHESES: 1) E2 reduces Par-4 expression and modifies its activity as well as its localization, abrogating its ability to induce apoptosis in endometrial cells. Therefore, inhibition of E2 activity should reinstate Par-4 expression and activity. This hypothesis will be explored in Obj1. 2) Par-4 is negatively regulated through the non-genomic E2 pathway or through PI3/Akt axis; this pathway would also influence Par-4 localization through TGFß signaling, which plays a pivotal role in cell survival and EMT/MET. This hypothesis will be explored in Obj2. The overall and long-term objective of the proposed research program is to examine and understand the regulation of expression as well as the endocrine, paracrine and autocrine activity of Par-4 in the regulation of endometrial cell fate. EXPERIMENTAL APPROACHES: Known estrogen responsive and non-responsive endometrial cells will be used for in vitro studies. Mouse pregnancy and induced decidualization will be used to study TGF-ß and Par-4 axis in the EMT process. SPECIFIC OBJECTIVES: 1) To determine the role of E2, through ERa and PI-3K/Akt pathway, in the control of Par-4 expression, localization and activity; 2) To elucidate how Par-4 differentially induce apoptosis and EMT and the key mechanisms governing these processes. INNOVATION & SIGNIFICANCE: A deeper understanding of E2 and TGFß-dependent Par-4 regulation is essential in uterine functions. Information gained from these studies will also help to identify the molecular mechanisms controlling apoptosis/survival and other types of cell fate (migration, invasion, EMT/MET) and will provide important clues in our understanding of reproductive functions.