Physiological significance of G protein gamma subunit heterogeneity

异三聚体G蛋白在细胞表面受体到细胞内效应器的信号转导中起着重要作用。p和y亚基在生理条件下紧密结合，到目前为止，已经发现了11种γ亚型。为了阐明伽马亚基异质性的生理意义，我们搜索了伽马亚型之间的差异，得到了以下结果。1. 在成纤维细胞中，γ - 12亚基与f -肌动蛋白相关，而γ - 5定位于局灶粘连，这表明两种γ亚型的功能差异。2. 确定蛋白激酶C对γ - 12的磷酸化位点为Ser-2.3。γ - 12在细胞内受佛波酯、血清、溶血磷脂酸和生长因子等多种试剂的刺激而发生磷酸化。4. 为了检查γ亚基的作用，在NIH 3T3细胞中，各种γ亚型与β 1亚基一起过表达。γ - 12的表达诱导细胞圆角和细胞迁移增加，而其他γ亚基的表达未引起显著变化。5. 删除n端区域或用谷氨酸取代γ - 12的Ser-2消除了γ - 12的这些影响，而用谷氨酸取代Ser-2的突变体显示出与野生型γ - 12相当的影响。这些结果表明，γ - 12 Ser-2位点的磷酸化增强了细胞的运动性。6. 免疫组织化学研究表明，gamma3定位于神经元细胞，而gamma12定位于脑内的一些胶质细胞。7. 我们鉴定了从牛肺中纯化的betagamma5复合物制备中的beta4蛋白。β 4被证明选择性地与γ - 5和γ - 12相互作用。

Heterotrimeric G proteins play a major role in signal transduction from cell-surface receptors to intracellular effectors. The p and y subunits are tightly bound under physiological conditions, and so far, 11 isoforms of gamma have been found. To elucidate the physiological significance of gamma subunit heterogeneity, we searched the differences among gamma isoforms and obtained following results. 1. In fibroblasts, the gamma12 subunit associated with F-actin, while the gamma5 localized at focal adhesions, suggesting functional differences for the two gamma isoforms. 2. The phosphorylation site of gamma12 by protein kinase C was determined to be Ser-2.3. The gamma12 was phosphorylated in the cells by the stimulation with various reagents, such as phorbol ester, serum, lysophosphatidic acid and growth factors. 4. To examine the roles of gamma subunits, various gamma isoforms were overexpressed in NIH 3T3 cells together with the beta1 subunit. Expression of gamma12 induced cell rounding and increase of cell migration, but expression of other gamma subunits did not induce significant changes. 5. Deletion of the N-terminal region or replacement by alanine of Ser-2 of gamma12 eliminated these effects of gamma12, while a mutant in which Ser-2 was replaced with glutamic acid showed effects equivalent to wild-type gamma12. These results indicated that phosphorylation of gamma12 at Ser-2 enhances the motility of cells. 6. Immunohistochemical studies showed that gamma3 was localized in neuronal cells, while gamma12 was localized some glial cells in the brain. 7. We identified the beta4 protein in the preparation of betagamma5 complex purified from bovine lung. The beta4 was shown to interact selectively with gamma5 and gamma12.