Studies on the role of the distal sequences involved in the expression of the human growth hormone/chorionic somatomammotropin gene family

The human growth hormone/chorionic somatomammotropin (GH/CS) gene family offers an interesting model to study mechanisms involved in tissue-specific transcriptional regulation of genes. Despite their high sequence homology, the members of GH/CS gene family are predominantly expressed in two different tissues: the GH-N gene is expressed in pituitary, whereas the CS-L, CS-A, GH-V and CS-B genes are expressed in placenta. A series of highly conserved regions within the GH/CS gene locus were identified by other investigators and were suggested to play a role in the differential and tissue-specific transcriptional regulation of the GH/CS genes. Presented here is the characterization of sequences/elements residing within the GH/CS locus, as well as those found in the flanking regions. Previous work in our laboratory identified PSF (PSF-A/-B) elements which are involved in the pituitary repressor mechanism for the CS/GH-V genes. Part of the present thesis work is aimed at the identification of the PSF-DNA binding proteins using conventional protein purification methods combi ed with PSF-specific DNA affinity chromatography. Characterization of partially purified proteins using southwestern blotting suggested that PSF proteins could be in the molecular range of 50-70 kDa. Further analysis of the PSF-DNA sequence revealed that these elements are recognized likely by nuclear factor-1 (NF-1) or NF-1-like proteins. NF-1 belongs to a family of proteins whose molecular sizes range from 50 to 70 kDa. Thus, these characteristics strongly suggest that PSFs could be NF-1 or NF-1-like. An AP-1 element is identified in the upstream sequences of the GH/CS genes using Nase I protection assays. Attempts were also made to characterize the role of this AP-I element through the use of transient transcription analysis. The enhancer/enhancer-like sequences located downstream of CS-L, CS-A and CS-genes were shown to contain multiple elements for transcriptional enhancer factor-1 (TEF-1). The presence of these sites implicated TEF-1, which ...