Wang H Y, Watkins D C, Malbon C C
Department of Biochemistry, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Nature. 1992 Jul 23;358(6384):334-7. doi: 10.1038/358334a0.
Fully-differentiated mouse 3T3-L1 fibroblasts accumulate large amounts of lipid at 7-10 days after induction by insulin or by dexamethasone and a methyl xanthine. G proteins mediate transmembrane signalling from a diverse group of cell-surface receptors to effector units that include phospholipase C, adenylyl cyclase and ion channels. They are also targets of regulation themselves. 3T3-L1 fibroblasts display marked changes in levels of G protein when induced to differentiate to adipocytes. Here we show that cholera toxin, which ADP-ribosylates and activates the G protein subunit Gs alpha, blocks the induction of differentiation, whereas increasing intracellular cyclic AMP directly with the dibutyryl analogue or indirectly with pertussis toxin or forskolin does not affect differentiation. Oligodeoxynucleotides antisense to the sequence encoding Gs alpha accelerate differentiation markedly. The time course of adipogenesis declined from 7-10 days in controls to roughly 3 days in cultures treated with antisense-Gs alpha oligodeoxynucleotides, whereas oligodeoxynucleotides, antisense to Gi alpha 1, Gi alpha 3, and sense and missense to Gs alpha, had no such effect. Antisense-Gs alpha alone induced differentiation by day 7, indicating that Gs alpha activity modulates differentiation in 3T3-L1 cells, acting in a new role which is independent of increased intracellular cAMP.
完全分化的小鼠3T3-L1成纤维细胞在胰岛素、地塞米松和甲基黄嘌呤诱导后7-10天会积累大量脂质。G蛋白介导从多种细胞表面受体到效应器单元的跨膜信号传导，这些效应器单元包括磷脂酶C、腺苷酸环化酶和离子通道。它们自身也是调节的靶点。当3T3-L1成纤维细胞被诱导分化为脂肪细胞时，其G蛋白水平会发生显著变化。在这里我们表明，霍乱毒素可使G蛋白亚基Gsα发生ADP核糖基化并激活它，从而阻断分化诱导，而用二丁酰类似物直接增加细胞内环状AMP或用百日咳毒素或福斯高林间接增加细胞内环状AMP则不影响分化。与编码Gsα的序列反义的寡脱氧核苷酸能显著加速分化。脂肪生成的时间进程从对照中的7-10天缩短至用反义Gsα寡脱氧核苷酸处理的培养物中的约3天，而与Giα1、Giα3反义的寡脱氧核苷酸以及与Gsα正义和错义的寡脱氧核苷酸则没有这种作用。单独的反义Gsα在第7天就诱导了分化，这表明Gsα活性调节3T3-L1细胞的分化，其作用方式独立于细胞内环状AMP的增加，发挥着新的作用。
