The human growth hormone gene family is composed of six genes coding for three proteins of physiological and clinical importance: growth hormone (hGH), prolactin (hPrl) and chorionic somatomammotropin (hCS). The correct and regulated expression of these genes is important for maintaining normal growth and development, starting from embryonic life through adulthood. Five of the genes are located on human chromosome 17 within a 60 Kb cluster, where the gene order is: hGH-N, hCS-A, hGH-V and hCS-B, but the sixth gene, hPrl, is located on chromosome 6. The high degree of sequence homology (less than 93%) and clustering suggest that the hGH and hCS genes evolved recently, probably by gene amplification. In spite of their similarity and close proximity, the expression of these genes is strictly tissue specific. hGH-N coding for GH is expressed in the anterior pituitary, while the other four genes, coding for various forms of CS and a rare variant of GH, are expressed in the placenta. On the other hand, the more distantly related Prl gene is also expressed in the anterior pituitary. While the complete structure and chromosomal organization of these genes are known, the mechanisms responsible for their strict tissue specific expression are not understood. We propose to study these mechanisms first by defining the various cis-acting genetic elements responsible for the remarkable differential expression of the hGH-N and hCS-A genes and second by characterizing the trans-acting regulatory factors which recognize these cis-elements. Once such factors are found, they will be purified to near homogeneity and their biological significance will be tested in vivo and in vitro by a variety of genetic and biochemical approaches. Finally, partial amino-acid sequences of the factors, which are of primary importance for conferring tissue specific expression upon these genes, will be determined. In addition, specific antisera will be raised against these factors and together, with oligonucleotide probes which match the protein sequence, will be used for screening of cDNA libraries. We also plan to develop a new approach for detecting clones coding for DNA binding proteins, by screening expression libraries with synthetic oligonucleotide probes which match the recognition sites of the various DNA binding proteins and a genetic selection procedure that will allow direct selection of the gene coding for one of these trans acting factors. Once the appropriate cDNA clones are isolated, they will be used for isolating the regulatory genes which control the expression of the GH gene family. We hope that studying the structure, function and regulation of the regulatory genes will reveal some of the molecular mechanisms which underlie tissue specific gene-expression.
Showing the most recent 10 out of 17 publications
