RhoA Signaling and Stroke

出血性卒中占所有卒中病例的约13%。这种类型的主要潜在案例之一 中风是颅内动脉瘤（IA）破裂，随后发生蛛网膜下腔出血， 受影响血管下游的血流减少。家族性和散发性IA病例的最新研究 在欧洲人和亚洲人中，发现ArhGEF 17基因突变与IA风险增加相关 人口。ArhGEF 17编码RhoA特异性的Dbl家族GEF（鸟嘌呤核苷酸交换因子）。的 ArhGEF 17的分子结构对于Dbl-family GEF来说是不寻常的，因为预计它具有不寻常的 DH结构域下游的分裂PH结构域（sPH）和预测的β-推进器（WD 40）结构域。有趣的是， 人类基因组有两个具有这种结构的ArhGEF 17同源物：ArhGEF 10和GRINCH-GEF （ArhGEF 10 L）（参见图1）。ArhGEF 10 L基因的多态性也与风险增加有关。 缺血性中风这三种蛋白质的结构尚不清楚，它们的具体功能尚不清楚。 推测的sPH和WD 40结构域尚不清楚。 我们提出了一个探索性的项目，旨在结构和功能表征的ArhGEF 17/10。我们 将描述ArhGEF 17/10家族的结构特征，其总体目标是获得 了解这些蛋白质的结构-功能关系。我们的最终目标是获得详细的 了解三个球状域的结构和功能，并确定是否以及如何 由超模块化结构调节的GEF。围绕ArhGEF 17的生理功能，我们 想要确定它是否通过下调其在平滑肌中的表达来调节脑血管张力， 小鼠大脑中动脉和内皮细胞中，使用腺病毒shRNA。RhoA活性和血管舒张 将在基础条件下以及对管腔内压力和激动剂增加的反应中进行监测， 动脉内皮细胞中的膜电阻测量将监测渗透性的变化。这 将为将来开发条件性ArhGEF 17敲除小鼠奠定基础，以确定 是否在平滑肌或内皮细胞（或两者）中缺失ArhGEF 17导致脑血管生成增加， 血管扩张和/或内皮通透性增加，并增加脑出血的倾向。

Hemorrhagic stroke accounts for ~13% of all stroke cases. One of the main underlying cases of this type of stroke is the rupture of intracranial aneurysm (IA), with consequent subarachnoid hemorrhage and critically reduced blood flow downstream of the affected vessel. A recent study of both familial and sporadic IA cases identified mutations in the ArhGEF17 gene as associated with increased risk for IA in both European and Asian populations. ArhGEF17 codes for a Dbl-family GEF (guanine nucleotide exchange factor) specific for RhoA. The molecular architecture of ArhGEF17 is unusual for a Dbl-family GEF, in that it is predicted to harbor an unusual split PH-domain (sPH) downstream of the DH domain, and a predicted β-propeller (WD40) domain. Interestingly, the human genome has two homologues of ArhGEF17 with this architecture: ArhGEF10 and GRINCH-GEF (ArhGEF10L) (see Fig 1). A polymorphism in the ArhGEF10L gene has also been associated with increased risk to ischemic stroke. The structures of these three proteins are not known, and the specific functions of the putative sPH and WD40 domains are not understood. We propose an exploratory project aimed at structural and functional characterization of ArhGEF17/10. We will characterize the structural features of the ArhGEF17/10 family, with the overarching goal to gain understanding of the structure-function relationships in these proteins. Our ultimate objective is to gain detailed understanding of the structure and functions of the three globular domains, and to determine if and how are the GEFs regulated by the supramodular architecture. Focusing on the physiological function of ArhGEF17, we want to determine if it regulates cerebral vascular tone, by down regulating its expression in smooth muscle in mouse mid cerebral arteries, and in endothelial cells, using adenoviral shRNAs. RhoA activity and vasodilation will be monitored under basal conditions and in response to increases in intraluminal pressure and agonists in the arteries. Membrane resistance measurements in endothelial cells will monitor changes in permeability. This will lay the ground work for the future development of a conditional ArhGEF17 knockout mouse to determine whether deletion of ArhGEF17 in either smooth muscle or endothelial cells (or both) leads to increased cerebral vessel dilation and/or increased endothelial permeability, and increases the propensity for cerebral hemorrhage.