Ca2 Second Messenger for Control of Flagellar Movement

钙离子作为第二信使在调控细胞凋亡中的作用 将检查鞭毛搏动。 建议的主要目标 实验的目的是利用生物化学方法， 鞭毛轴丝中的结合蛋白可能参与 控制鞭毛弯曲的形状。 生化分级 从莱茵衣藻中分离的轴丝将进行， 确定钙调素（CaM）的亚轴丝分布， 与鞭毛跳动的控制有关 的分布 将通过凝胶覆盖监测轴丝中的钙调素结合蛋白 使用125-I-CaM或生物素化CaM和抗生物素蛋白-过氧化物酶的方法。 将从轴丝制备外部双重微管以确定 无论这些结构是否含有CaM、CaM结合蛋白或其他 感兴趣的钙结合蛋白。 最后，多克隆抗血清 将制备针对特异性CaM结合蛋白的抗体， 用于免疫标记研究，以检查这种 “候选”蛋白质，位于轴丝和外部双联体之间 微管 这些研究应该阐明钙改变 鞭毛波形通过识别关键的钙结合蛋白， 控制鞭毛的形状，以及一般扩大我们的理解， 钙作为第二信使的过程。

The role of calcium ions as a second messenger in the control of flagellar beat will be examined. The major objective of the proposed experiments is to identify, using biochemical procedures, calcium- binding proteins in the flagellar axoneme that may be involved in controlling the shape of flagellar bends. Biochemical fractionation of axonemes isolated from Chlamydomonas reinhardtii will be carried out to determine the sub-axonemal distribution of calmodulin (CaM), which has been implicated in the control of flagellar beat. The distribution of CaM-binding proteins in the axoneme will be monitored by gel overlay procedures using 125-I-CaM or biotinylated CaM and avidin-peroxidase. Outer doublet microtubules will be prepared from axonemes to determine whether these structures contain CaM, CaM-binding proteins, or other calcium binding proteins of interest. Finally, polyclonal antisera directed against a specific CaM-binding protein will be prepared and used in immunolabelling studies to examine the distribution of this "candidate" protein wthin the axoneme and the outer doublet microtubules. These studies should clarify the mechanisms by which calcium alters flagellar waveform by identifying key calcium-binding proteins that may control flagellar shape, as well as generally broaden our understanding of processes that involve calcium as a second messenger.