The fully differentiated vascular smooth muscle cell (SMC) is endowed with a genetic program of growth cessation and cell-restricted contractile gene expression, both of which are compromised in vascular disease. The molecular mechanisms that underlie SMC contractile gone down-regulation likely involve perturbations in the expression and/or binding activity of transcription factors recognizing multiple regulatory modules. Such modules may exist hundreds of kilobases from the core promoter making their identification difficult by conventional transgenesis. Indeed, for years investigators have been stymied by the complete lack of promoter activity in transgenic mice with one particularly recalcitrant SMC-restricted gene, smooth muscle calponin (SM-Calp). The advent of bacterial artificial chromosomes (BACs), however, has facilitated the inclusion of all regulatory modules controlling expression of human SM-Calp in transgenic mice. Emerging technologies in the field of bioinformatics have accelerated SM-Calp regulatory module discovery in BACs, including our preliminary identification of an intergenic boundary element that appears to function as an insulator. These data, along with the recent discovery of a novel cell-restricted transcription factor, form the basis of this competitive renewal application, which seeks to test the hypothesis that SM-Calp requires multiple evolutionarily-conserved regulatory modules and binding factors whose functional inactivity accounts for reduced expression of SM-Calp in vascular disease. An innovative, multidisciplinary approach involving bioinformatics, traditional wet-lab assays, BAC transgenesis, and in vivo models of arterial injury is planned to map and define all regulatory modules controlling SM-Calp expression in vivo. These studies will provide the necessary groundwork for elucidating the transcriptional circuitry involved with the down-regulation of SM-Calp. Such information has enormous value for understanding and potentially treating vascular diseases as well as pathological vessel development.
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