Regulation of axonemal Mg2+-ATPase from Paramecium cilia: effects of Ca2+ and cyclic nucleotides.
Ciliary activity is regulated by Ca2+ and cyclic nucleotides, but the molecular mechanisms of the regulation are unknown. We have tested the ability of Ca2+ and cyclic nucleotides to alter ciliary Mg2+-ATPase or to stimulate phosphorylation of axonemal dynein. Mg2+-ATPase activity in cilia and axonemes from Paramecium was stimulated 2-fold by micromolar Ca2+, but this Ca2+ sensitivity was lost upon solubilization of the dyneins from the axoneme. The Ca2+-sensitive component of ciliary Mg2+-ATPase activity was inhibited by the dynein inhibitors vanadate and Zn2+, but was insensitive to the calmodulin antagonists calmidazolium and melittin. Dynein activity in the high-salt extract from axonemes was also insensitive to calmidazolium. Calmodulin did not sediment with 22 S or 12 S dyneins on sucrose gradients containing Ca2+, but it did sediment in the region from 19 S to 14 S. Mg2+-ATPase activity in ciliary fractions was unaltered in the presence of cAMP or cGMP. However, polypeptides associated with the 22 S and 12 S dyneins, as well as proteins of 19 S, 15 S, and 8 S, were substrates for endogenous ciliary kinases. High molecular weight polypeptides that sedimented at 22 S and 19 S were phosphorylated in a cyclic nucleotide-stimulated manner.
Travis
S M
SM
Department of Biochemistry, College of Agriculture and Life Sciences, University of Wisconsin-Madison 53706.
Nelson
D L
DL
D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D002923:Cilia / Q000378:metabolism; D009712:Nucleotides, Cyclic / Q000494:pharmacology*; D010247:Paramecium / Q000187:drug effects / Q000378:metabolism*; D010766:Phosphorylation
D002147:Calmodulin; D009712:Nucleotides, Cyclic; D017301:Ca(2+) Mg(2+)-ATPase; D002118:Calcium