It has become clear that as neuroectodermal stem cells evaginate from the oral primordia and migrate superiorly to join peptidergic neurons of the posterior pituitary, that transcription activators and repressors are likely to interact in order to establish various and discrete cell types that comprise the anterior pituitary gland. Our understanding of the molecular details which dictate the tissue-restricted expression of pituitary-specific genes has been advanced by the discovery of pit-1/GHF- 1, a pituitary-specific transcription factor which contains a POU- homeodomain and regulates the promoter activity of the growth hormone (GH), prolactin (PRL), and Pit-1/GHF-1 genes. Pit-1 serves not only as a transcription activator of these genes, but it also plays a critical role as a differentiation factor, specifying the ontogeny of the somatotroph, lactotroph, and thyrotroph pituitary cell lineages. Nevertheless, the presence or absence of a tissue-specific transcription activator, such as Pit-1/GHF-1, cannot account for the 7 to 8 orders of magnitude of differential GH and PRL gene expression between pituitary and nonpituitary tissues. The achievement of such high levels of tissue-restricted gene expression implies that repressors of GH and PRL gene expression must be present in nonpituitary cell types. We have discovered the presence of a repressor of rPRL promoter activity in nonpituitary cell types, and of a ubiquitous trans-activator required for basal rPRL promoter activity, and we have mapped the rPRL DNA elements to which they bind. The repressor, termed F2F, binds to the footprint (FP) II site, and it appears to functionally interfere with the basal activating function of the vicinally-bound BTF, which binds to the basal transcription element, or BTE. Our data suggests a model whereby the inhibitory effect of F2F is dependent on the BTE DNA context and, therefore, the DNA-binding of BTF. Furthermore, these factors appear to modulate the hormonal response of the rPRL promoter; most likely by influencing Pit-1 bound to the most proximal cell-specific element.
The Specific Aims we propose in order to test this model directly, are: (1) to fully characterize the F2F/FP II and BTF/BTE protein DNA interactions; (2) to functionally dissect the FP II and BTE DNA elements; (3) to determine mechanism by which F2F and/or BTF modulate hormonal control of the rPRL promoter; (4) to purify F2F and BTF; and (5) to clone cDNAs for F2F and BTF. The significance of these studies is that they will fully elucidate the precise molecular mechanism(s) by which ubiquitous factors govern the activity of a highly tissue-restricted gene. Since there is little understanding of the fundamental processes by which ubiquitous factors influence tissue-specific gene expression, these studies are very likely to provide novel insights into the molecular mechanisms by which the basal transcription machinery governs tissue- specific gene expression.
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