Physiological significance of G protein gamma subunit heterogeneity

异源三聚体G蛋白在从细胞表面受体到细胞内效应物的信号转导中起主要作用。p和y亚基在生理条件下紧密结合，迄今已发现11种γ亚基异构体，为了阐明γ亚基异质性的生理意义，我们寻找γ异构体之间的差异，并获得以下结果。1.在成纤维细胞中，gamma 12亚基与F-肌动蛋白相关，而gamma 5定位于局灶性粘连，表明两种γ亚型的功能差异。2.蛋白激酶C对γ 12的磷酸化位点被确定为Ser-2.3。用佛波酯、血清、溶血磷脂酸和生长因子刺激细胞，γ 12被磷酸化。4.为了研究γ亚基的作用，各种γ亚型与β 1亚基一起在NIH 3 T3细胞中过表达。γ 12的表达诱导细胞变圆和细胞迁移的增加，但其他γ亚基的表达没有引起显着的变化。5.γ 12的N-末端区域的缺失或Ser-2的丙氨酸取代消除了γ 12的这些作用，而Ser-2被谷氨酸取代的突变体显示出与野生型γ 12相当的作用。这些结果表明，γ 12在Ser-2的磷酸化增强了细胞的运动性。6.免疫组织化学研究表明，γ 3定位于神经元细胞，而γ 12定位于脑中的一些胶质细胞。7.我们鉴定了从牛肺中纯化的β-γ 5复合物的制备中的β 4蛋白。β 4被证明可以选择性地与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.