Localization of Calmodulin and Dynein Light Chain Lc8 in Flagellar Radial Spokes

doi:10.1083/jcb.153.6.1315

Published online 11 June 2001. doi:10.1083/jcb.153.6.1315

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© The Rockefeller University Press, 0021-9525/2001//1315 $5.00
The Journal of Cell Biology, Volume 153, Number 6, , 2001 1315-1326

Original Article

Localization of Calmodulin and Dynein Light Chain Lc8 in Flagellar Radial Spokes

Pinfen Yang^a, Dennis R. Diener^b, Joel L. Rosenbaum^b, and Winfield S. Sale^a

^a Department of Cell Biology, Emory University, School of Medicine, Atlanta, Georgia 30322
^b Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, Connecticut 06520
Department of Cell Biology, Emory University School of Medicine, 1648 Pierce Dr., Atlanta, GA 30222.(404) 727-6256(404) 727-6265

win{at}cellbio.emory.edu

Genetic and in vitro analyses have revealed that radial spokesplay a crucial role in regulation of ciliary and flagellar motility,including control of waveform. However, the mechanisms of regulationare not understood. Here, we developed a novel procedure toisolate intact radial spokes as a step toward understandingthe mechanism by which these complexes regulate dynein activity.The isolated radial spokes sediment as 20S complexes that arethe size and shape of radial spokes. Extracted radial spokesrescue radial spoke structure when reconstituted with isolatedaxonemes derived from the radial spoke mutant pf14. Isolatedradial spokes are composed of the 17 previously defined spokeproteins as well as at least five additional proteins includingcalmodulin and the ubiquitous dynein light chain LC8. Analysesof flagellar mutants and chemical cross-linking studies demonstratedcalmodulin and LC8 form a complex located in the radial spokestalk. We postulate that calmodulin, located in the radial spokestalk, plays a role in calcium control of flagellar bending.

Key Words: calcium • cilia • dynein • flagella • calmodulin

Abbreviations used in this paper: 2D, two-dimensional; KI, potassiumiodide; KP, potassium phosphate; pI, isoelectric point; PKA,protein kinase A.

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