Morton D B, Truman J W
Department of Zoology, University of Washington, Seattle 98195.
J Neurosci. 1988 Apr;8(4):1326-37. doi: 10.1523/JNEUROSCI.08-04-01326.1988.
The present study describes 2 phosphoproteins, both with an apparent molecular weight of 54 kDa, in the CNS of the tobacco hornworm, Manduca sexta. Their phosphorylation is regulated by a neuropeptide, eclosion hormone (EH), and the second messenger cGMP, which thus have been named the EGPs (eclosion hormone- and cGMP-regulated phosphoproteins). Although cAMP was more effective than cGMP at stimulating the phosphorylation of the EGPs in CNS homogenates, in the intact CNS cGMP was more effective. Since cGMP mediates the action of EH, this strongly suggests that cGMP is the second messenger that stimulates the phosphorylation of the EGPs in vivo. The EGPs can only be phosphorylated in vitro during discrete time periods during the life of Manduca. During the larval and pupal molts, the EGPs can first be phosphorylated just prior to ecdysis. Their ability to be phosphorylated is correlated with the time when the insect is sensitive to EH. This close temporal correlation suggests that the ability to phosphorylate the EGPs determines when the insect can first respond to EH. During adult development, the EGPs first appeared on fluorograms 6 d before sensitivity to EH, suggesting that at this stage other factors may also be involved in the regulation of sensitivity. For the ecdyses of all 3 stages, EH appeared to stimulate the phosphorylation of the EGPs at ecdysis. The EGPs were found in all regions of the prepupal nervous system that were investigated, but only in the abdominal and pterothoracic ganglia of the developing adult. Fractionation of nervous system homogenates by ultracentrifugation revealed that one of the EGPs was present only in the pellet fraction, whereas the other was approximately equally distributed between pellet and supernatant. Furthermore, the EGPs in the pellet fraction could be partially solubilized with detergents and high salt concentrations.
本研究描述了烟草天蛾（Manduca sexta）中枢神经系统中的两种磷蛋白，二者的表观分子量均为54 kDa。它们的磷酸化受一种神经肽——羽化激素（EH）以及第二信使环鸟苷酸（cGMP）的调节，因此被命名为EGP（羽化激素和cGMP调节的磷蛋白）。尽管在中枢神经系统匀浆中，环磷酸腺苷（cAMP）比cGMP更有效地刺激EGP的磷酸化，但在完整的中枢神经系统中，cGMP更有效。由于cGMP介导EH的作用，这强烈表明cGMP是在体内刺激EGP磷酸化的第二信使。EGP仅在烟草天蛾生命周期中的特定时间段内才能在体外被磷酸化。在幼虫和蛹蜕皮期间，EGP可在蜕皮前首次被磷酸化。它们被磷酸化的能力与昆虫对EH敏感的时间相关。这种紧密的时间相关性表明，EGP被磷酸化的能力决定了昆虫何时能首次对EH作出反应。在成虫发育过程中，EGP首次出现在对EH敏感前6天的荧光图谱上，这表明在此阶段其他因素可能也参与了敏感性的调节。对于所有三个阶段的蜕皮，EH似乎在蜕皮时刺激EGP的磷酸化。在所研究的蛹前期神经系统的所有区域均发现了EGP，但仅在发育中的成虫的腹部和翅胸神经节中发现。通过超速离心对神经系统匀浆进行分级分离显示，其中一种EGP仅存在于沉淀部分，而另一种在沉淀和上清液中分布大致相等。此外，沉淀部分中的EGP可用去污剂和高盐浓度部分溶解。
