Regulation of flagellar dynein activity by a central pair kinesin

doi:10.1073/pnas.0406817101

Regulation of flagellar dynein activity by a central pair kinesin

^*Department of Cell and Developmental Biology, State University of New York Upstate Medical University, 750 East Adams Street, Syracuse, NY 13210; and ^‡Boulder Laboratory for 3-D Electron Microscopy of Cells, Department of Molecular, Cellular, and Developmental Biology, University of Colorado, Boulder, CO 80309-0347

Edited by J. Richard McIntosh, University of Colorado, Boulder, CO, and approved October 22, 2004 (received for review September 14, 2004)

Abstract

The motility of cilia and flagella is powered by dynein ATPases associated with outer doublet microtubules. However, a flagellar kinesin-like protein that may function as a motor associates with the central pair complex. We determined that Chlamydomonas reinhardtii central pair kinesin Klp1 is a phosphoprotein and, like conventional kinesins, binds to microtubules in vitro in the presence of adenosine 5′-[β,γ-imido]triphosphate, but not ATP. To characterize the function of Klp1, we generated RNA interference expression constructs that reduce in vivo flagellar Klp1 levels. Klp1 knockdown cells have flagella that either beat very slowly or are paralyzed. EM image averages show disruption of two structures associated with the C2 central pair microtubule, C2b and C2c. Greatest density is lost from part of projection C2c, which is in a position to interact with doublet-associated radial spokes. Klp1 therefore retains properties of a motor protein and is essential for normal flagellar motility. We hypothesize that Klp1 acts as a conformational switch to signal spoke-dependent control of dynein activity.

Footnotes

↵ ¶ To whom correspondence should be addressed. E-mail: mitcheld{at}upstate.edu.
↵ † Present address: Department of Biology, Emory University, Atlanta, GA 30322.
↵ § Deceased October 21, 2001.
Author contributions: R.Y. and D.R.M. designed research; R.Y., S.G., and D.R.M. performed research; R.Y., E.O., and D.R.M. analyzed data; and D.R.M. wrote the paper.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: IFT, intraflagellar transport; RNAi, RNA interference; AMP-PNP, adenosine 5′-[β,γ-imido]triphosphate.
↵ ∥ Dymek, E. E., Wargo, M. J. & Smith, E. F. (2003) Mol. Biol. Cell 14, 433a (abstr.).