Hepatic GPR125: ligands and liver repair

项目摘要 G蛋白偶联受体(GPCRs)是7种跨膜区蛋白，它们有助于维持 通过对内部和外部环境产生的信号作出反应而实现的动态平衡。在人类中，有 超过800种已知的GPCR，虽然FDA批准的所有药物中有超过20%针对GPCR，但大量的 这些受体中的大多数都没有得到充分的研究。事实上，许多这些未确定特征的“孤儿”受体， 被称为GPR的蛋白在不同的组织中表达水平极高，这表明它们可能 重要但未知的生理功能。肝脏是人体内最大的代谢组织 并负责一些重要的功能，包括感应和解毒外源生物， 代谢葡萄糖和其他能源，合成和分泌胆汁酸。因此，这是很好的- 准备利用一些感官GPR来帮助这些过程。我们最近确认了总共57个 GPR在小鼠肝脏中不同程度地表达，包括7个表达谱与之竞争的GPR 其他特性良好的受体。其中一个高表达的受体是黏附受体GPR125 (ADGRA3)。作为一个家族，黏附受体与维持细胞-细胞连接和 已被报道通过蛋白水解性裂解其非共价连接的N-末端而激活 域。然而，GPR125是一种带有突变切割位点的非典型黏附受体，目前尚不清楚 这种受体是通过传统的过程激活的。GPR125已被认为与发展 斑马鱼模型中的平面细胞极性提示该受体可能参与成年的组织修复 纸巾。利用RNAScope和免疫荧光技术，我们已经将GPR125定位于基底膜 胆管细胞。胆管内的胆管细胞在肝再生中起着关键作用。 受伤。因此，GPR125特别适合为这一进程做出贡献。我们假设GPR125，在 胆管细胞的基侧膜，由细胞外基质的成分激活，以便 有助于损伤后的肝脏修复。这将使用Presto-Tango实验进行测试，以使其去孤儿 CRISPR导向型腺病毒载体介导的受体和独特的肝特异性基因敲除 RNA。总而言之，我们的发现将揭示一种高度表达但未被充分描述的， 并将为未来在小鼠和人身上进行生理分析奠定基础。

Project Summary G protein-coupled receptors (GPCRs) are 7 transmembrane domain proteins that aid in the maintenance of homeostasis by responding to cues produced by the internal and external environment. In humans, there are more than 800 known GPCRs, and while more than 20% of all FDA-approved drugs target GPCRs, the vast majority of these receptors are understudied. In fact, many of these uncharacterized ‘orphan’ receptors, termed GPRs, are expressed at extremely high levels in different tissues suggesting that they likely have important, yet unknown, physiological functions. The liver is the largest metabolic tissue in the human body and is responsible for a number of important functions including sensing and detoxifying xenobiotics, metabolizing glucose and other energy sources, and synthesizing and secreting bile acids. Thus, it is well- poised to take advantage of some sensory GPRs to aid in these processes. We recently identified a total of 57 GPRs that are expressed to varying degrees in the murine liver, including 7 with expression profiles that rival other well characterized receptors. One of these high expressing receptors is adhesion receptor GPR125 (ADGRA3). As a family, adhesion receptors have been linked to the maintenance of cell-cell junctions and have been reported to be activated by proteolytically cleavage of their non-covalently attached N-terminal domain. However, GPR125 is an atypical adhesion receptor with a mutated cleavage site and it is unclear if this receptor is activated via the conventional processes. GPR125 has been linked to the development of planar cell polarity in a Zebrafish model suggesting that this receptor may be involved in tissue repair in adult tissues. Using RNAscope and immunofluorescence, we have localized GPR125 to the basolateral membrane of cholangiocytes. Lining the bile ducts, cholangiocytes play a key role in the liver regeneration following injury. Thus, GPR125 is uniquely suited to contribute to this process. We hypothesize that GPR125, on the basolateral membrane of cholangiocytes, is activated by components of the extracellular matrix in order to contribute to liver repair following injury. This will be tested using the PRESTO-TANGO assay to deorphanize the receptor alongside a unique liver-specific knockout mediated by adenoviral delivery of CRISPR guide RNAs. Collectively, our discoveries will unveil new functions for a highly expressing, yet under-characterized, receptor and will set the stage for future physiological analysis in both mouse and man.