Growth hormone is critical for growth and development. Its gene is expressed specifically in the pituitary and has been intensely studied as a model to understand hormone action, growth control, and gene expression. We have studied this gene and have identified several factors that interact with its promoter. High level promoter expression requires combinatorial actions of several factors that are both tissue-specific and more ubiquitous. Thus, it is critical to understand how these factors work. The current studies focus on a factor, GHF-3, that contributes substantially to GH expression; it exerts a ca. 4-8-fold influence, comparable to that elicited through other DNA sites. GHF-3 may be a novel factor that is more generally involved in regulating gene expression, since it is abundant in multiple tissues and the DNA sequence that it binds appears to be unique. GHF-3 forms multimeric complexes with other proteins that may reflect a potential for complex regulatory influences. Finally, GHF-3 can be readily studied, as its complexes with other proteins and DNA binding properties can be demonstrated, it can be purified, and its activity can be shown by gene transfer and in in vitro transcription systems. In the proposed studies, it is planned to define the DNA site that binds GHF-3 to determine if it is present in other promoters. GHF-3 cDNA will be cloned by either expression screening with radiolebelled GHF-3 site DNA or by purifying GHF-3 and using its sequence to prepare primers. The cDNA sequence should provide information about the factor and its relatedness to other transcriptional regulators. The cDNA will be used to screen tissues for GHF-3 mRNA and to express GHF-3 in microbes, mammalian cells and/or cell free systems. The expressed protein, or GHF-3 purified from liver, will be used to prepare antiserum to GHF-3 that will be used to probe tissues for GHF-3 expression, characterize potential variant forms and post-translational modifications of GHF-3, and better define GHF-3 interactions with other proteins. Vectors expressing GHF-3 cDNA will be used in gene transfer experiments to understand GHF-3 action, and mutated cDNAs will be used to define functional domains of GHF-3 required for activity and interactions with other factors. Finally, GHF-3 function will be studied in an in vitro transcription system. The information derived should further our knowledge of control of growth hormone gene expression and transcription control in general.
| Schaufele, F; West, B L; Baxter, J D (1992) Synergistic activation of the rat growth hormone promoter by Pit-1 and the thyroid hormone receptor. Mol Endocrinol 6:656-65 |
