It is well known that cellular 90-kilodalton heat shock protein (HSP-90) can complex with steroid receptors including estrogen receptors (ER) and modify their DNA and steroid binding properties. In female reproductive tissues, the cellular level of HSP-90 itself is regulated by estradiol (E2) which can also modulate the growth of these tissues and some of their tumors. HSP-90 belongs to a subset of HSPs which are developmentally regulated in a wide variety of species and alterations in the cellular regulation of HSP-90 are commonly associated with transformed cells. Accordingly, we believe that in female reproductive tissues E2 and HSP-90 may mutually regulate their cellular functions to maintain cellular replicative homeostasis. Therefore, the long-term objectives of this proposal is to elucidate the potential regulatory role of HSP-0 in E2- dependent growth in normal and neoplastic female reproductive tissues. In mammalian cells, HSP-90 is encoded by two distinct genes, HSP-84 and 86 and are speculated to have different functions which also respond to E2 with different magnitudes. Therefore, our immediate objective is to resolve the cellular and molecular basis for E2-dependent synthesis of HSP-84 and 86 using the well defined rodent uterus as a model system.
The specific aims are: 1) To examine the effect of E2 on HSP-84 and 86 mRNA and protein in the various uterine cell types by performing immunocytochemical analyses for the protein and in situ hybridization for mRNA, using antibodies and cDNA probes unique to each gene product. 2) To identify the DNA elements on murine HSP-84 and 86 genes responsible for mediating E2-dependent expression and examine if E2 and ER modulate the interaction of these elements with their corresponding cellular factors. For this, we will isolate and sequence the promoter and 5' flanking regions of HSP-84 and 86 genes, test these for their E2 inducibility, and also examine their interactions with cellular extracts isolated from control and E2 treated tissues. In all these experiments, the involvement of ER will also be examined. Since alterations in the cellular levels of HSPs are believed to have important consequences with regard to the outcome of certain treatment modalities such as radiation, hyperthermia and chemotherapy. Information obtained from these studies can have a major significance in evaluating the outcome of certain treatment modalities used for cancers of the female reproductive tract. It may also help to structure these treatment protocols with respect to their selection, sequences and time intervals for administration.
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