Agonist-independent, or constitutive, activity is an integral feature of G protein-coupled receptors, but its relevance in pathophysiological settings is generally poorly explored. GPR35 is a therapeutic target in inflammatory diseases of the lower gut. In colonic organoids from a human GPR35a-expressing transgenic mouse line, the GPR35 inverse agonist CID-2745687 increased barrier permeability substantially, indicating that constitutive receptor activity contributes to maintaining epithelial barrier integrity. High constitutive activity of GPR35 was also observed in both HT-29 and HEPG2 cells that express GPR35 endogenously. Mechanistic investigations in recombinant in vitro systems revealed that the constitutive activity of GPR35a was biased and not equivalent across signaling pathways. Hence, no constitutive interactions of the receptor with arrestin-adaptor proteins or activation of Gαo-containing G protein heterotrimers were detected while, even at low GPR35a expression levels, substantial constitutive activation of heterotrimers containing either Gα12 or Gα13 was observed. Similar biased constitutive activity was observed for the human GPR35b isoform. The extent of constitutive and agonist-mediated activity was dependent on receptor expression level. At high receptor levels, constitutive activation of Gα12 or Gα13 masked any agonist-induced effects while low expression levels with low constitutive activity allowed measurement of agonist-induced responses. These results highlight roles, selectivity, and the extent of constitutive activity of GPR35 in cells and tissues that express this receptor endogenously and highlight the contribution of its constitutive activity to maintaining the colonic epithelial barrier, potentially limiting the development of inflammatory bowel diseases.

中文翻译：

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G 蛋白偶联受体 GPR35 的偏倚组成型信号传导抑制肠道屏障通透性


激动剂非依赖性或组成型活性是 G 蛋白偶联受体的一个组成部分特征，但其在病理生理学环境中的相关性通常很少得到探索。GPR35 是下肠道炎症性疾病的治疗靶点。在表达 GPR35a 的人转基因小鼠系的结肠类器官中，GPR35 反向激动剂 CID-2745687 显着增加了屏障通透性，表明组成型受体活性有助于维持上皮屏障完整性。在内源性表达 GPR35 的 HT-29 和 HEPG2 细胞中也观察到 GPR35 的高组成型活性。重组体外系统的机制研究表明 ，GPR35a 的组成型活性是偏倚的，并且在信号通路中不等效。因此，未检测到受体与抑制蛋白接头蛋白的组成性相互作用或含有 Gαo 的 G 蛋白异源三聚体的激活，而即使在低 GPR35a 表达水平下，也观察到含有 Gα12 或 Gα13 的异源三聚体的实质性组成型激活。在人 GPR35b 亚型中观察到类似的偏倚组成型活性。组成型和激动剂介导的活性程度取决于受体表达水平。在高受体水平下，Gα12 或 Gα13 的组成型激活掩盖了任何激动剂诱导的作用，而具有低组成型活性的低表达水平允许测量激动剂诱导的反应。 这些结果突出了 GPR35 在内源性表达该受体的细胞和组织中的作用、选择性和组成活性程度，并强调了其组成型活性对维持结肠上皮屏障的贡献，可能限制炎症性肠病的发展。