The Characteristic Laboratory of Crop Germplasm Innovation and Application, Provincial Department of Education, College of Agronomy, Qingdao Agricultural University, Qingdao 266109, China.
State Key Laboratory of Crop Biology, College of Life Sciences, Shandong Agricultural University, Tai'an 271018, China.
Plant Physiol. 2023 Aug 3;192(4):3170-3188. doi: 10.1093/plphys/kiad238.
Heterotrimeric GTP-binding proteins (G proteins) are a group of regulators essential for signal transmission into cells. Regulator of G protein signaling 1 (AtRGS1) possesses intrinsic GTPase-accelerating protein (GAP) activity and could suppress G protein and glucose signal transduction in Arabidopsis (Arabidopsis thaliana). However, how AtRGS1 activity is regulated is poorly understood. Here, we identified a knockout mutant of oxysterol binding protein-related protein 2A, orp2a-1, which exhibits similar phenotypes to the arabidopsis g-protein beta 1-2 (agb1-2) mutant. Transgenic lines overexpressing ORP2A displayed short hypocotyls, a hypersensitive response to sugar, and lower intracellular AtRGS1 levels than the control. Consistently, ORP2A interacted with AtRGS1 in vitro and in vivo. Tissue-specific expression of 2 ORP2A alternative splicing isoforms implied functions in controlling organ size and shape. Bioinformatic data and phenotypes of orp2a-1, agb1-2, and the orp2a-1 agb1-2 double mutant revealed the genetic interactions between ORP2A and Gβ in the regulation of G protein signaling and sugar response. Both alternative protein isoforms of ORP2A localized in the endoplasmic reticulum (ER), plasma membrane (PM), and ER-PM contact sites and interacted with vesicle-associated membrane protein-associated protein 27-1 (VAP27-1) in vivo and in vitro through their two phenylalanines in an acidic track-like motif. ORP2A also displayed differential phosphatidyl phosphoinositide binding activity mediated by the pleckstrin homology domain in vitro. Taken together, the Arabidopsis membrane protein ORP2A interacts with AtRGS1 and VAP27-1 to positively regulate G protein and sugar signaling by facilitating AtRGS1 degradation.
异三聚体 G 蛋白结合蛋白（G 蛋白）是一组对于信号传入细胞至关重要的调节剂。G 蛋白信号调节因子 1（AtRGS1）具有内在的 GTP 酶加速蛋白（GAP）活性，可抑制拟南芥（Arabidopsis thaliana）中的 G 蛋白和葡萄糖信号转导。然而，AtRGS1 活性如何被调节还知之甚少。在这里，我们鉴定了一个氧化固醇结合蛋白相关蛋白 2A（或 orp2a-1）的敲除突变体，该突变体表现出与拟南芥 G 蛋白β 1-2（agb1-2）突变体相似的表型。过表达 ORP2A 的转基因系表现出短的下胚轴、对糖的超敏反应和比对照更低的细胞内 AtRGS1 水平。一致地，ORP2A 在体外和体内与 AtRGS1 相互作用。2 个 ORP2A 选择性剪接异构体的组织特异性表达暗示了控制器官大小和形状的功能。orp2a-1、agb1-2 和 orp2a-1 agb1-2 双突变体的生物信息学数据和表型揭示了 ORP2A 和 Gβ 在 G 蛋白信号和糖反应调节中的遗传相互作用。ORP2A 的这两种选择性蛋白异构体都定位于内质网（ER）、质膜（PM）和 ER-PM 接触位点，并通过其酸性轨道样模体中的两个苯丙氨酸与囊泡相关膜蛋白相关蛋白 27-1（VAP27-1）在体内和体外相互作用。ORP2A 还在体外显示出由 PH 结构域介导的不同的磷酯酰磷酸肌醇结合活性。总之，拟南芥膜蛋白 ORP2A 与 AtRGS1 和 VAP27-1 相互作用，通过促进 AtRGS1 降解来正向调节 G 蛋白和糖信号。
