Microtubule-membrane interactions in cilia: II. photochemical cross-linking of bridge structures and the identification of a membrane-associated dynein-like ATPase

Abstract

Photochemical cross-linking of both Tetrahymena and Aequipecten ciliary membrane proteins with the lipophilic reagent 4,4′-dithiobisphenylazide links together a high molecular weight dynein-like ATPase, membrane tubulin, and at least two other proteins. Electron microscopy of detergent-extracted cilia reveals that the cross-linked complex remains attached to the outer-doublet microtubules by a microtubule-membrane bridge. Cleavage of the reagent’s disulfide bond releases the bridge-membrane complex and the dynein-like membrane-associated ATPase. Electron microscopy was used to ensure that the dynein-like protein did not result from the solubilization of the dynein arms attached to the outer-doublet microtubules. The dynein-like protein has been isolated using sucrose gradients and is similar to axonemal dynein with respect to its sedimentation characteristics, nucleotide specificity, and divalent cation requirements. Photochemical crosslinking of ciliary membrane proteins in vivo results initially in the modification of ciliary beat and, eventually, in the cessation of ciliary movement. These results suggest that a dynein-like ATPase comprises the bridge which links the ciliary membrane to the outer-doublet microtubles and that this bridge is involved in the modulation of normal ciliary movement. © 1980, Rockefeller University Press., All rights reserved.