Microtubule-membrane interactions in cilia. II. Photochemical cross-linking of bridge structures and the identification of a membrane-associated dynein-like ATPase.
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 cross-linking of ciliary membrane porteins 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 microtubules and that this bridge is involved in the modulation of normal ciliary movement.
D000251:Adenosine Triphosphatases / Q000032:analysis*; D000818:Animals; D001386:Azides; D002462:Cell Membrane / Q000201:enzymology / Q000648:ultrastructure*; D002923:Cilia / Q000502:physiology / Q000648:ultrastructure*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D008565:Membrane Proteins / Q000032:analysis; D008870:Microtubules / Q000648:ultrastructure*; D008974:Mollusca / Q000502:physiology / Q000648:ultrastructure*; D009068:Movement; D012995:Solubility; D013501:Surface-Active Agents; D013769:Tetrahymena pyriformis / Q000502:physiology / Q000648:ultrastructure*; D014466:Ultraviolet Rays
D001386:Azides; D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D013501:Surface-Active Agents; C025369:4,4'-dithiobis(phenylazide); D000251:Adenosine Triphosphatases