ANABOLIC HORMONES OF DOMINANT NEGATIVE REGULATION

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Primates of the New World (i.e. South and Central America) or platyrrhines are resistant to the cellular actions of the gonadal steroid hormone 17beta-estradiol. Over the span of the last 50 million years platyrrhines have evolved to accommodate this divergence from the hormone-responsive phenotype characteristic of the only other surviving primate suborder, Catarrhini or Old World primates, which include our own species Homo sapiens. Our central hypothesis states that this hormone-resistant phenotype represents the incorporation of a successful gain-of-function mutation by primordial New World primates that resulted in the constitutive overexpression of a family of dominant- negative-acting hormone response element binding proteins in the heterogeneous ribonucleoprotein (hnRNP) superfamily of nucleic acid binding proteins. Two corollaries to our hypothesis hold that this gain-of-function mutation 1] was designed to interrupt estrogen-directed signal transduction independent of the receptor proteins for the hormone, and 2] has been refined over millions of years to require the cooperative action of an hsp-70-related subfamily of high-capacity, intracellular ligand binding proteins. The purpose of this application is to explore the biochemical basis for this extraordinary and successful experiment of Nature by investigating prototypical members of these two novel families of primate signaling proteins, the estrogen-response element binding (ERE-BP) and the intracellular estrogen binding protein (IEBP). The first specific aim is to dissect the molecular basis for the regulated overexpression of the ERE-BP in platyrrhine cells. This goal will be achieved by the application of state-of-the-art technology in 1] tissue- and cell-specific localization of ERE-BP mRNA and protein, 2] functional analysis of the ERE-BP promoter on both an estrogen-resistant and wild-type background, and 3] targeted overexpression of the ERE-BP in estrogen-responsive tissues (i.e. breast and bone). The second specific aim is to 1] purify IEBP, 2] clone its full- length cDNA, and 3] transiently overexpress that cDNA to determine how IEBP co-legislates estrogen responsiveness in platyrrhines. Because there are Old World primate homologs of these overexpressed New World primate proteins, the ultimate aim of this research program is to employ Nature's as well as our own transgenic experiment and to understand the roles of these dominant-negative-acting proteins in estrogen-modified human health and disease (i.e. osteoporosis, breast cancer).