The genes of the human growth hormone/chorionic somatomammotropin (GH/CS) gene cluster are expressed specifically in syncytiotrophoblastic epithelium of the fetal placenta (hCS-A, hCS-B, hCS-L, and hGH-V) or in somatotrope and somatolactotrope cells of the anterior pituitary (hGH-N). The overall aims of this research program are to define the basis for the high-level, tissue-specific, developmentally-controlled expression of this gene cluster in vivo, to define the full spectrum of encoded protein products, and to delineate the form and function of the cognate receptors that mediate the tissue response to the encoded hormones.
The SPECIFIC AIMS of this proposal focus on: 1) Defining how the GH/CS gene locus is organized at the level of chromatin structure, the mechanism and ontogeny of its activation via its locus control region (LCR), the organization and function of its LCR and flanking regulatory elements, and how they result in placenta- versus pituitary-specific expression. As a corollary, a survey of GH-deficient human patients may determine whether genetic mutations of the LCR can result in human disease; 2) Establishing novel pituitary cell lines that can serve as critical reagents for the definition and characterization of the LCR, by targeted immortalization of somatotrope cells in mice transgenic for the GH locus and LCR; 3) Establishing a transgenic mouse model in which the entire GH/CS gene cluster with its LCR and flanking regulatory components can be studied in detail in their native configuration in vivo; 4) Characterizing the structure and function of the alternatively-spliced, exon 3 deficient hGH receptor isoform normally expressed in placental villi, and its potential role in placental growth and development; and 5) Characterizing the expression of the alternatively-processed transcripts and protein products of the two least characterized genes in the cluster: hGH-V and hCS-L. The methods used to achieve these aims include standard techniques of molecular biology, transgenic mouse technology including targeted oncogenesis, detection of clones in yeast artificial chromosome libraries and their analysis and expression, cell culture and transfection, microinjection of Xenopus oocytes, and protein analysis with gels and antisera. The significance of this line of investigation will be the detailed understanding of the expression and action of the GH/CS cluster and its regulation as a unit by the LCR. This will serve as a foundation for a sophisticated understanding of the pathophysiology of human growth and development, and may lead in turn to the design of new means of medical intervention.
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