Delineating the role of CTF and NTF of Gpr126 in physiology and pathophysiology

粘附G蛋白偶联受体（agpcr）信号传导的主要激活机制是隐性系链激动剂与7个跨膜结构域的相互作用。这种激动剂可以在GAINdomain内的GPS基序上进行自动催化裂解，产生n端片段（NTF）和c端片段（CTF），或者在NTF上发生变化（如配体结合或机械力）时通过异构化释放。aGPCRGpr126 （Adgrg6）对心脏、耳部和雪旺细胞的发育至关重要。然而，Gpr126敲除（KO）小鼠表现出心脏和周围神经表型，而缺乏cttf的斑马鱼突变体仅表现出周围神经表型，这表明entf在心脏中具有特定作用。此外，斑马鱼的救援实验表明，GPR126具有区域特异性功能。然而，这种现象在任何其他aGPCR中都没有被描述过，在小鼠中也没有数据可用，并且受体切割是否重要以及单个结构域具有哪些功能仍然不清楚。为了证明NTF具有独立于CTF的功能，我们在第一个资助期获得或生成了几种动物模型，包括Gpr126报告小鼠、条件Gpr126 KO小鼠和几种转基因斑马鱼系。对这些模型的分析确定了gpr126表达改变后的几种大体形态学以及细胞和分子表型。在第二个资助期内，我们将继续对Gpr126进行结构-功能分析。我们工作的一个重要焦点是确定Gpr126 KO小鼠的胚胎致死表型是否确实是由于心脏发育缺陷引起的，以及心内膜细胞中ntf的表达是否足以促进心脏发育。此外，我们将阐明Gpr126在心脏瓣膜发育中的作用，因为我们的数据表明Gpr126在内皮间质转化中起作用。最后，我们将描述Gpr126在肾脏生理和病理生理中的作用。我们的数据表明Gpr126在几种肾细胞类型中表达。此外，它的表达在损伤后高度上调。最后，我们的数据表明NTF可能在心脏和肾脏中以旁分泌的方式起作用。总的来说，我们已经产生了各种各样的工具，这些工具将使我们能够描述Gpr126的CTF和ntf在生理学和病理生理学中的作用，有助于更好地理解aGPCR信号传导和功能。

The major activation mechanism of adhesion G protein-coupledreceptor (aGPCRs) signaling is the interaction of a cryptic tetheredagonist with the seven transmembrane domain. This agonist can beliberated by autocatalytical cleavage at the GPS motif within the GAINdomain, yielding an N-terminal fragment (NTF) and a C-terminalfragment (CTF) or through isomerization upon changes occurring atthe NTF such as ligand binding or mechanical force. The aGPCRGpr126 (Adgrg6) is essential for heart, ear, and Schwann celldevelopment. However, while Gpr126 knockout (KO) mice exhibit aheart and peripheral nerve phenotype, zebrafish mutants lacking theCTF exhibit only the peripheral nerve phenotype, suggesting that theNTF has a specific role in heart. Moreover, rescue experiments inzebrafish suggest that GPR126 has domain-specific functions. Yet,such a phenomenon has not been described for any other aGPCR, nodata in mouse are available, and it remains unclear whether receptorcleavage is important as well as which functions the individualdomains have. To demonstrate that the NTF has an independentfunction from the CTF, we have obtained or generated during the firstfunding period several animal models including Gpr126 reporter mice,conditional Gpr126 KO mice and several transgenic zebrafish lines.Analysis of these models identified several gross morphological aswell as cellular and molecular phenotypes upon alteration of Gpr126expression. During the second funding period we will continue thestructure-function analysis of Gpr126. An important focus of our workis to determine if the embryonic lethal phenotype in Gpr126 KO miceis indeed due to defective heart development and whether NTFexpression in endocardial cells is sufficient for proper heartdevelopment. Further, we will elucidate the role of Gpr126 in heartvalve development as our data suggest a role for Gpr126 inendothelial mesenchymal transformation. Finally, we will characterizethe role of Gpr126 during kidney physiology and pathophysiology. Ourdata indicate that Gpr126 is expressed in several renal cell types.Moreover, its expression is highly upregulated upon injury. Finally, ourdata suggest that the NTF might act in the heart as well as in thekidney in a paracrine manner. Collectively, we have generated a widevariety of tools that will allow us to delineate the role of CTF and NTFof Gpr126 in physiology and pathophysiology contributing to a betterunderstanding of aGPCR signaling and function.