whirlin蛋白与espin蛋白相互作用在视网膜色素变性发病机制中的研究

Retinitis Pigmentosa （RP） is a degenerative eye disease caused by abnormalities of the photoreceptors or the retinal pigment epithelium of the retina. Usher syndrome is a disease that combines RP and hearing loss. It is classified into three clinical types, with Type II being the most common form accounting for about 70% of all cases. Genetic defects in three genes (whirlin, ush2a and vlgr-1) are known to underlie this type of Usher syndrome. This complex of the products from these three genes is localized mainly at the periciliary membrane complex in photoreceptors and the ankle-link of the stereocilia in hair cells. Whirlin can recruit the other two proteins. In our previous study, we identified Espin, an actin-binding / bundling protein involved in human deafness when defective, as a whirlin-interacting protein. The interaction between these two proteins was confirmed by their coimmunoprecipitation and colocalization in cultured cells. This interaction involves multiple domains of both proteins and only occurs when Espin does not bind to actin. However, the biological function of this USH2 protein complex is largely unknown. . In this study, we will use biochemistry, genetic, co-transfection, Co-immunoprecipitation(CoIP) and in vivo tools and methods to further study the functions of these proteins and their protein complex. Our study will further identify and explore the significance of the interaction between Whirlin and Espin. This study can provide insights on the mechanisms how defects in USH2 complex may lead to Usher syndrome. This study will provide insights on developing novel therapeutic approaches for this group of diseases.

视网膜色素变性(retinitis pigmentosa, RP),是一组以进行性感光细胞及色素上皮功能丧失为共同表现的慢性进行性退行性病变，Usher综合征II型(USH2)就是以RP为特征的疾病。Whirlin,ush2a,vlgr-1是USH2的主要致病基因。它们在视网膜感光细胞的连接纤毛（connecting cilium,CC）处组成蛋白复合体。CC结构的异常是导致RP发病的主要原因。在我们前期工作中，已经发现并证实Espin是USH2蛋白复合物的一个新成员，且和 Whirlin存在相互作用。但是，这两个蛋白的具体相互作用以及蛋白复合体的生物功能还不是很清楚。我们将通过对这两个蛋白的进一步研究，以及用whirlin的N/C末端的质粒和espin全长的质粒共转染细胞，用免疫共沉淀的方法发现这两个蛋白的具体结合位点，对探索RP的发病机制及今后基因治疗、新药开发提供一个崭新的视角

Usher综合征是一种以失明和失聪为特点的严重性疾病。已经证实有三个基因被确认为Usher综合征II的致病基因，分别为whirlin,ush2a,vlgr-1称为Usher 复合体。我们以往实验证实了espin是Usher 复合体的一个新成员，whirlin和espin的相互作用对疾病的发生起到重要的作用，whirlin还可以调节和espin相结合的actin的移动性。然而，whirlin的哪个区域和espin存在相互作用还很不清楚，从而阻碍对疾病的理解和治疗的进一步发展。. whirlin有三个PDZ区域和一个脯氨酸聚集区域,这些都是蛋白质和蛋白质相互作用的区域。众所周知，whirlin是一个支架蛋白，在组成USH2蛋白质复合体中起决定性作用。这里Whirin-N末端亚型包括全长的第一、二个PDZ区域和PR区域，whirin-C端亚型包括第三个PDZ区域。荧光共聚焦定位实验证明whirlin的N末端和espin全长有共定位，免疫共沉淀实验证实这俩个蛋白存在相互作用。. 在这里,我们从蛋白相互作用的多角度证实whirlin的N末端和espin存在相互作用，对疾病的发生发展发病机制有进一步的理解。这对视网膜色素变性后续的基因诊断及靶向治疗提供了重要的依据和方向。