Ekstrom R C, Hunzicker-Dunn M
Department of Cell, Molecular, and Structural Biology, Northwestern University Medical School, Chicago, Illinois 60611.
Endocrinology. 1990 Nov;127(5):2578-86. doi: 10.1210/endo-127-5-2578.
Pig ovarian follicular membranes contain a gonadotropin-responsive adenylyl cyclase, which becomes partially desensitized (approximately 40%) upon a 40-min incubation with a saturating concentration of human (h) CG. This in vitro desensitization is time and hormone dependent and also requires the presence of micromolar concentrations of GTP. In this report we show that 10% ethanol present during the desensitization phase of the incubation increases the extent of hCG-induced desensitization of adenylyl cyclase by 2-fold. Ethanol shortened the time necessary to reach maximal hCG-induced desensitization from 20 to 10 min, but had no effect on the dose dependency for GTP. In addition, ethanol had no effect on the affinity of the LH/hCG receptor for 125I-hCG but did cause an increase in the ED50 of hCG for inducing desensitization from 0.25 to 0.75 nM. Interestingly, ethanol decreased the apparent number of LH/hCG-receptor sites by 55%, yet the control hCG-sensitive adenylyl cyclase activity was not reduced. The "hyperdesensitized" state achieved in the presence of ethanol could not be reversed by washing the membranes and incubating them in ethanol-free medium. NaF-sensitive adenylyl cyclase was also not impaired in hCG-desensitized membranes from control or ethanol-treated samples. Thus, hCG-induced desensitization was not due to a defect in the functioning of the stimulatory guanine nucleotide-binding regulatory protein (G8) or catalytic subunits, but rather was caused by an impairment of the coupling of the lutropin (LH)/hCG receptor with G8, which was exacerbated further by ethanol. In spite of the effect of ethanol on hCG-induced desensitization, this agent had an inhibitory effect on isoproterenol-induced desensitization of isoproterenol-responsive luteal adenylyl cyclase. These results indicate that membrane fluidity is important in modulating the structure and functional interaction of the LH/hCG receptor with G8 because ethanol is a well known lipid-fluidizing agent. The resistance to ethanol potentiation of desensitization of the isoproterenol-sensitive adenylyl cyclase suggests that there are differences between the LH/hCG- and beta-adrenergic receptors in factors controlling their structures and or interactions with G proteins, and that there is a fundamental difference in their mechanisms of desensitization.
猪卵巢卵泡膜含有一种促性腺激素反应性腺苷酸环化酶，在与饱和浓度的人绒毛膜促性腺激素（hCG）孵育40分钟后，该酶会部分脱敏（约40%）。这种体外脱敏是时间和激素依赖性的，并且还需要微摩尔浓度的鸟苷三磷酸（GTP）存在。在本报告中，我们表明在孵育的脱敏阶段存在的10%乙醇可使hCG诱导的腺苷酸环化酶脱敏程度增加2倍。乙醇将达到最大hCG诱导脱敏所需的时间从20分钟缩短至10分钟，但对GTP的剂量依赖性没有影响。此外，乙醇对促黄体生成素（LH）/hCG受体与125I-hCG的亲和力没有影响，但确实导致hCG诱导脱敏的半数有效剂量（ED50）从0.25 nM增加到0.75 nM。有趣的是，乙醇使LH/hCG受体位点的表观数量减少了55%，然而对照hCG敏感的腺苷酸环化酶活性并未降低。在乙醇存在下达到的“超敏”状态不能通过洗涤膜并在无乙醇培养基中孵育来逆转。在对照或乙醇处理样品的hCG脱敏膜中，氟化钠敏感的腺苷酸环化酶也未受损。因此，hCG诱导的脱敏不是由于刺激性鸟嘌呤核苷酸结合调节蛋白（G8）或催化亚基功能缺陷，而是由促黄体素（LH）/hCG受体与G8的偶联受损引起的，乙醇进一步加剧了这种情况。尽管乙醇对hCG诱导的脱敏有影响，但该试剂对异丙肾上腺素诱导的异丙肾上腺素反应性黄体腺苷酸环化酶脱敏有抑制作用。这些结果表明膜流动性在调节LH/hCG受体与G8的结构和功能相互作用中很重要，因为乙醇是一种众所周知的脂质流化剂。异丙肾上腺素敏感的腺苷酸环化酶对乙醇增强脱敏的抗性表明，在控制其结构和/或与G蛋白相互作用的因素方面，LH/hCG受体和β-肾上腺素能受体之间存在差异，并且它们的脱敏机制存在根本差异。
