The capacity of vascular smooth muscle cells (SMCs) to modulate their response to arterial injury has been implicated in the pathogenesis of vascular proliferative syndromes including atherosclerosis and restenosis following coronary intervention. We have utilized the SMC-specific mouse SM22a promoter as a model system to elucidate the transcriptional programs that control VSMC development and differentiation. During the initial funding cycle of this award, the cis-acting elements and trans-acting factors that control expression of the SM22a promoter were defined. Two nuclear protein binding sites in the SM22a promoter (SME- 1 and SME-4) were identified which contain consensus CArU boxes that bind specifically to the MADS box transcription factor, SRF. Remarkably, a multimerized copy of either SME-l or SME-4 is necessary and sufficient to restrict activity of a LacZ reporter gene to arterial SMCs in transgenic mice. These data lead us to hypothesize that SRF, in concert with other potentially novel transcriptional activators (and potentially repressors), regulates SMC-specific gene expression and SMC phenotype. The overall goal of the proposed studies is to elucidate the molecular basis of SRF-dependent transcription in arterial SMCs.
The specific aims of these studies are to: i) Examine the molecular basis of SRF-CArG box activity in SMCs, ii) Examine protein-protein associations that modulate activity of the SM22a promoter in SMCs with particular attention on SRF/homeobox protein interactions, iii) Examine signaling pathways underlying LIMK- 1-mediated activation of the SM22cz promoter, and iv) Examine the capacity of SRF-/- ES cells to contribute to the SMC lineage(s) in SRF-/- -C57BL/6 chimenc mice and define downstream SMC and mesodermal genes regulated by SRF in the embryo. Taken together, these studies will elucidate the transcriptional program and an important signaling pathway that modulates expression of the SM22a gene in SMCs. Because CArG box-containing elements have been identified in multiple other SMC-specific genes these studies will provide fundamental insights into the molecular mechanisms that control SMC differentiation and the modulation of vascular SMC phenotype.
