Zhang Y H, Ross E M, Snell W J
Department of Cell Biology and Neuroscience, University of Texas Southwestern Medical School, Dallas 75235.
J Biol Chem. 1991 Dec 5;266(34):22954-9.
Adenylylcyclase activity in the flagella of gametes of Chlamydomonas reinhardtii was inhibited by prior incubation at or below 30 degrees C in the presence of ATP. This decrease did not occur in the absence of ATP, in the presence of the ATP analog 5'-adenylylimidodiphosphate (App(NH)p), or in the presence of ATP plus the protein kinase inhibitor staurosporine (2 microM). If ATP treatment was performed in the absence of an ATP-regenerating system, activity initially declined and subsequently recovered. Incubation of flagella at 45 degrees C in the absence of ATP or incubation at lower temperatures in the presence of either App(NH)p or staurosporine both increased adenylylcyclase activity (over 10-fold) and blocked subsequent ATP-dependent loss of activity at 30 degrees C. This heat-induced activation was prevented by the presence of ATP plus an ATP-regenerating system. Incubation of flagella with [gamma-32P]ATP followed by gel electrophoresis in sodium dodecyl sulfate indicated the presence of endogenous protein kinase and protein phosphatase activities. These data suggest that the flagellar adenylylcyclase in Chlamydomonas gametes is inhibited by phosphorylation and stimulated by dephosphorylation. This mechanism for regulating adenylylcyclase may underlie the rapid increase in cyclic AMP that is induced by flagellar adhesion during fertilization in Chlamydomonas.
莱茵衣藻配子鞭毛中的腺苷酸环化酶活性在30摄氏度及以下、ATP存在的条件下预先孵育后受到抑制。在没有ATP、存在ATP类似物5'-腺苷酰亚胺二磷酸（App(NH)p）或存在ATP加蛋白激酶抑制剂星形孢菌素（2 microM）的情况下，这种活性降低不会发生。如果在没有ATP再生系统的情况下进行ATP处理，活性最初会下降，随后恢复。在没有ATP的情况下将鞭毛在45摄氏度孵育，或在存在App(NH)p或星形孢菌素的情况下在较低温度孵育，都会增加腺苷酸环化酶活性（超过10倍），并阻止随后在30摄氏度时ATP依赖性的活性丧失。ATP加ATP再生系统的存在可阻止这种热诱导的激活。用[γ-32P]ATP孵育鞭毛，然后在十二烷基硫酸钠中进行凝胶电泳，表明存在内源性蛋白激酶和蛋白磷酸酶活性。这些数据表明，莱茵衣藻配子中的鞭毛腺苷酸环化酶受到磷酸化抑制，去磷酸化则会刺激其活性。这种调节腺苷酸环化酶的机制可能是衣藻受精过程中鞭毛黏附诱导环磷酸腺苷快速增加的基础。
