VLGR1 signaling at focal adhesions and ICD release as a non-canonical signaling pathway of aGPCRs

VLGR1 (very large G protein偶联受体-1)，又称adgrv1、GPR98和MASS，是迄今为止最大的粘附性GPCR（aGPCR）。虽然VLGR1的表达几乎无处不在，但它高度集中于神经系统。人类vlgr1基因的突变导致人类Usher综合征（USH），这是遗传性聋盲的最常见形式。此外，越来越多的证据表明，VLGR1缺陷也与癫痫有关。迄今为止，对其在膜粘附和信号传导中的双重功能知之甚少。在目前的资助期内，我们通过应用亲和蛋白质组学（tandemaffinity纯化（TAPs）结合质谱）确定了与VLGR1和其他几种agpcr相关的相互作用伙伴和蛋白质簇。对这些aGPCR相互作用组的生物信息学分析已经定义了基因本体（GO）术语和与可能的aGPCR功能相关的细胞模块。在第二个资助期，我们选择了特定的功能模块来进一步评估和深入表征它们对VLGR1和agpcr的信号功能。(1)我们将阐明VLGR1在局灶黏附（FAs）中的作用，FAs是ECM和细胞生长和迁移所必需的肌动细胞骨架之间的信号枢纽。我们将使用多种VLGR1缺陷细胞模型（包括不同类型的神经细胞）来测试VLGR1对FAturnover的时空控制、细胞迁移的调节和细胞骨架的调节的贡献。(2)我们将破译vlgr1和其他aGPCR与γ -分泌酶复合物的相互作用，作为一种常见的非规范aGPCR信号通路。为此，我们将测试agpcr是否是γ -分泌酶的底物，以及它们的胞内结构域（ICDs）如何被释放到细胞质中。此外，我们将评估agpcr的核功能。我们确信我们的结果将有助于理解agpcr的工作原理。我们也希望获得与agpcr功能障碍相关的疾病的病理机制的新见解，并找到治疗这些疾病的新治疗靶点的机会。

VLGR1 (very large G protein-coupled receptor-1), also known asADGRV1, GPR98 and MASS is by far the largest adhesion GPCR(aGPCR). Although VLGR1 expression is almost ubiquitous, it ishighly concentrated in the nervous system. Mutations in the humanVLGR1 gene cause the human Usher syndrome (USH), the mostcommon form of hereditary deaf-blindness. In addition, there isgrowing evidence that defects in VLGR1 are also associated withepilepsy. So far, little is known about its dual function in membranemembraneadhesion and signaling. In the current funding period, weidentified interaction partners and protein clusters related to VLGR1,and to several other aGPCRs by applying affinity proteomics (tandemaffinity purifications (TAPs) in combination with mass spec).Bioinformatics analyses of the interactomes of these aGPCRs havedefined Gene Ontology (GO) terms and cell modules related toputative aGPCR function. For the second grant period, we haveselected specific functional modules for further evaluation and indepthcharacterization of their signaling function for VLGR1 and foraGPCRs. (1) We will elucidate the role of VLGR1 in focal adhesions(FAs) which act as signaling hubs between the ECM and the actincytoskeleton essential for cell outgrowth and migration. We will testVLGR1´s contribution to the spatial and temporal control of FAturnover, the regulation of cell migration and the modulation of thecytoskeleton using divers VLGR1 deficient cellular models includingdifferent types of neural cell. (2) We will decipher the interaction ofVLGR1 and other aGPCRs with the gamma-secretase complex as acommon non-canonical aGPCR signaling pathway. For this, we willtest whether aGPCRs are substrates of the gamma-secretase andhow their intracellular domains (ICDs) are released into thecytoplasm. Furthermore, we will evaluate nuclear functions ofaGPCRs. We are certain that our results will help to understand howaGPCRs work. We also hope to gain new insights into thepathomechanisms of diseases associated with the dysfunction ofaGPCRs and see opportunities to identify new therapeutic targets forthe treatment of these diseases.