非洲爪蟾卵母细胞中RC3/神经颗粒蛋白钙调蛋白结合结构域单一位点变体的功能研究

Functional studies of single-site variants in the calmodulin-binding domain of RC3/neurogranin in Xenopus oocytes.

作者信息

Watson J B, Margulies J E, Coulter P M, Gerendasy D D, Sutcliffe J G, Cohen R W

机构信息

Mental Retardation Research Center, UCLA School of Medicine 90024-1759, USA.

出版信息

Neurosci Lett. 1996 Nov 29;219(3):183-6. doi: 10.1016/s0304-3940(96)13203-1.

DOI:10.1016/s0304-3940(96)13203-1

PMID:8971810

Abstract

Single-site variants in the calmodulin-binding domain of RC3/neurogranin were heterologously expressed in Xenopus oocytes to examine their effects on serotonin-evoked currents. RC3 variants serine36 -->alanine (Ser36-->Ala), serine36-->glycine (Ser36-->Gly), and phenylalanine37-->tryptophan (Phe37-->Trp), which bind calmodulin but are deficient in protein kinase C (PKC) phosphorylation, display serotonin-evoked Ca(2+)-dependent Cl- currents in oocytes similar to control oocytes. A serine36-->aspartate (Ser36-->Asp) variant, which does not bind calmodulin and mimics the PKC-phosphorylated state of RC3, significantly enhances serotonin-evoked currents in a manner similar to wild-type. The results suggest that RC3 not only regulates the availability of free calmodulin in a dendritic spine but also, when phosphorylated, independently stimulates G-protein coupled second messenger pathways that generate inositol 1,4,5-trisphosphate (IP3), diacylglycerol (DAG) and intracellular Ca2+.

摘要

RC3/神经颗粒素钙调蛋白结合域中的单一位点变体在非洲爪蟾卵母细胞中进行异源表达，以研究它们对5-羟色胺诱发电流的影响。结合钙调蛋白但缺乏蛋白激酶C（PKC）磷酸化的RC3变体丝氨酸36→丙氨酸（Ser36→Ala）、丝氨酸36→甘氨酸（Ser36→Gly）和苯丙氨酸37→色氨酸（Phe37→Trp），在卵母细胞中显示出与对照卵母细胞相似的5-羟色胺诱发的Ca(2+)依赖性Cl-电流。不结合钙调蛋白且模拟RC3的PKC磷酸化状态的丝氨酸36→天冬氨酸（Ser36→Asp）变体，以类似于野生型的方式显著增强5-羟色胺诱发的电流。结果表明，RC3不仅调节树突棘中游离钙调蛋白的可用性，而且在磷酸化时，还独立刺激产生肌醇1,4,5-三磷酸（IP3）、二酰基甘油（DAG）和细胞内Ca2+的G蛋白偶联第二信使途径。