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J Cell Biol. 2009 Sep 21;186(6):817-24. doi: 10.1083/jcb.200906168. Epub 2009 Sep 14.

Regulation of dynein-driven microtubule sliding by the axonemal protein kinase CK1 in Chlamydomonas flagella.

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1
Department of Cell Biology, Emory University School of Medicine, Atlanta, GA 30322, USA.

Abstract

Experimental analysis of isolated ciliary/flagellar axonemes has implicated the protein kinase casein kinase I (CK1) in regulation of dynein. To test this hypothesis, we developed a novel in vitro reconstitution approach using purified recombinant Chlamydomonas reinhardtii CK1, together with CK1-depleted axonemes from the paralyzed flagellar mutant pf17, which is defective in radial spokes and impaired in dynein-driven microtubule sliding. The CK1 inhibitors (DRB and CK1-7) and solubilization of CK1 restored microtubule sliding in pf17 axonemes, which is consistent with an inhibitory role for CK1. The phosphatase inhibitor microcystin-LR blocked rescue of microtubule sliding, indicating that the axonemal phosphatases, required for rescue, were retained in the CK1-depleted axonemes. Reconstitution of depleted axonemes with purified, recombinant CK1 restored inhibition of microtubule sliding in a DRB- and CK1-7-sensitive manner. In contrast, a purified "kinase-dead" CK1 failed to restore inhibition. These results firmly establish that an axonemal CK1 regulates dynein activity and flagellar motility.

PMID:
19752022
PMCID:
PMC2753152
DOI:
10.1083/jcb.200906168
[Indexed for MEDLINE]
Free PMC Article
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