Localization and functional difference of various betagamma subunits of G protein

我们已经确定了从牛脾脏中纯化的G蛋白的一个新的γ亚基（gamma12，以前指定为gammaS1）的一级结构。由68个氨基酸组成的成熟γ - 12蛋白在n端有乙酰化丝氨酸，在c端有香叶酰化/羧化半胱氨酸。这与从牛脾脏cDNA文库中分离的gamma12 cDNA预测的氨基酸序列c端戊烯酰化信号一致。在各种γ亚基（γ 1, γ 2, γ 3， γ 7和γ 12）中，γ 12具有被蛋白激酶c磷酸化的独特性质，磷酸化的氨基酸残基是Ser1（或Ser2）。Swiss 3T3和主动脉平滑肌细胞暴露于phorbol 12-肉豆蔻酸酯13-乙酸烷NaF诱导γ - 12磷酸化。用血管紧张素和血管加压素等天然血管活性药物刺激主动脉平滑肌细胞也可诱导γ - 12的磷酸化。在体外，β - γ - 12的磷酸化程度被与α α形成的复合物所抑制，通过添加GTPgammaS或氟化铝来缓解。这些结果强烈表明，在活细胞中受体和G蛋白的激活过程中，γ - 12被蛋白激酶C磷酸化。然后，利用针对各自γ亚基的抗体，研究了G蛋白各种形式γ亚基的定位。γ - 3在神经细胞中表达，而γ - 12是大多数非神经细胞中主要的γ亚基。γ ma2、γ ma5和γ ma12分布在多种组织中，但γ ma2在大脑中占主导地位。

We have determined the primary structure of a novel gamma subunit (gamma12, previously designated gammaS1) of G protein purified from bovine spleen. The mature gamma12 protein composed of 68 amino acids had acetylated serine at the N-terminus and geranylgeranylated/carboxylmethylated cysteine at the Cterminus. This was consistent with the C-terminal prenylation signal in the amino acid sequence which was predicted from gamma12 cDNA isolated from a bovine spleen cDNA library. Among various gamma subunits (gamma1, gamma2, gamma3, gamma7 and gamma12), gamma12 has a unique property to be phosphorylated by protein kinase C.The phosphorylated amino acid residue was Ser1 (or Ser2). Exposure of Swiss 3T3 and aortic smooth muscle cells to phorbol 12-myristate 13-acetate ane NaF induced phosphorylation of gamma12.Stimulation of aortic smooth muscle cells with natural vasoactive agents such as angiotensin II and vasopressin also induced phosphorylation of gamma12. The extent of phosphorylation of betagamma12 in vitro was suppressed by a complex formation with Goalpha, which was relieved by the addition of GTPgammaS or aluminum fluoride. These results strongly suggest that gamma12 is phosphorylated by protein kinase C during activation of receptor (s) and G protein (s) in living cells. Then the localization of various forms of the gamma subunit of G protein was investigated by the use of antibodies against the respective gamma subunits. The gamma3 was expressed in neuronal cells, and gamma12 was the major gamma subunit in most nonneural cells. gamma2, gamma5 and gamma12 were distributed in a variety of tissues, but gamma2 was dominant in the brain.