Transcriptional regulation of CRP1 in smooth muscle

描述（由申请人提供）：血管平滑肌细胞是血管壁的重要组成部分，具有非凡的适应能力。这些细胞表现出一系列依赖于转录程序的选择性利用的表型。虽然平滑肌调节对于正常血管功能是必不可少的，但是它们的适应能力与血管闭塞疾病的病理学不利地相关。因此，基因转录的分子机制，导致产生表型的生物学和临床相关。作为阐明控制平滑肌转录的关键特征的进展，我们的研究集中在前体细胞向成熟平滑肌的发育和分化上。分化伴随着收缩所需的一组精确基因的精心激活。虽然确定性调控元件和相应的转录因子被认为在分化中发挥作用，但它们如何与传递选择性基因表达的信号偶联尚不清楚。我们假设细胞外信号，如转化生长因子-β（TGF-β），使用细胞内途径和转录因子的独特组合来赋予平滑肌限制性表达。TGF-β已经成为管理平滑肌细胞表型的主要候选者。该提议的目的是采用富含半胱氨酸的蛋白质1（CRP 1）基因的平滑肌特异性表达来研究顺式作用元件和反式作用因子，所述反式作用因子响应于限定的信号传导事件而传递选择性转录活性。我们已经确定了CRP 1基因的一个独特的调控区域，该区域仅在动脉平滑肌细胞中驱动表达。我们打算使用这种调控元件来表征CRP 1基因在分化的平滑肌细胞中表达的重要转录途径。具体目标是：1）确定转化生长因子β 1（TGF-β 1）激活CRP 1基因的转录机制。2)确定血清反应因子（SRF）的功能活性与控制平滑肌基因表达的信号事件之间的关系。3)明确确定CArG元件对通过靶向诱变表达CRP 1的必要性。这些研究意义重大，因为它们将超越转录调节因子的分析，并确定介导反式激活的确切途径，以解决有关平滑肌细胞表型表现的基本问题。

DESCRIPTION (provided by applicant): Vascular smooth muscle cells are a vital component of the blood vessel wall, possessing extraordinary adaptive abilities. These cells display a range of phenotypes that are dependent upon the selective utilization of transcriptional programs. While smooth muscle modulation is essential for normal blood vessel function, their adaptive abilities are adversely associated with the pathologies of vascular occlusion diseases. Consequently, the molecular mechanisms governing gene transcription leading to resultant phenotypes are both biologically and clinically relevant. As an inroad to elucidating critical features that control smooth muscle transcription, our studies are focused on the development and differentiation of precursor cells into mature smooth muscle. Differentiation is accompanied by the orchestrated activation of a precise set of genes required for contraction. Though definitive regulatory elements and corresponding transcription factors are recognized to have a role in differentiation, how they are coupled to signals that convey selective gene expression is not well understood. We hypothesize that extracellular signals, like transforming growth factor-beta (TGF-¿), use a distinct combination of intracellular pathways and transcription factors to impart smooth muscle-restricted expression. TGF-¿ has emerged as primary candidate for governing smooth muscle cell phenotypes. The aims of this proposal are designed to employ the smooth muscle-specific expression of the cysteine-rich protein 1 (CRP1) gene to investigate the cis-acting elements, and trans- acting factors that convey selective transcriptional activity in response to defined signaling events. We have identified a unique regulatory region of the CRP1 gene that drives expression exclusively in arterial smooth muscle cells. We intend to use this regulatory element to characterize essential transcriptional pathways important for the expression of the CRP1 gene in differentiated smooth muscle cells. The specific aims are: 1) To define the transcriptional mechanisms underlying the activation of the CRP1 gene by transforming growth factor-beta-1 (TGF-¿1). 2) To determine the relationship between the functional activity of serum response factor (SRF), and signaling events that govern smooth muscle gene expression. 3) To unequivocally determine the necessity of the CArG element for expression of CRP1 by targeted mutagenesis. These studies are significant, as they will extend beyond the analysis of the transcriptional regulators, and determine the exact pathways that mediate transactivation, to address fundamental questions regarding the manifestation of smooth muscle cell phenotypes.