Translocation and rotation of microtubules caused by multiple species of Chlamydomonas inner-arm dynein

Abstract

Dynein was extracted from outer arm-less axonemes of the mutant odal and fractionated by high-pressure liquid chromatography on a MonoQ column into seven distinct subspecies (named a–g). Each subspecies contained one or two heavy chains and several mediumsized and light chains; by vanadate/UV-induced photocleavage and SDS-polyacrylamide gel electrophoresis, eight distinct heavy chains were identified. Analysis of the mutant axonemes indicated that the subspecies f (containing two heavy chains) is missing in the inner-arm mutant idal and the subspecies a, c and d are missing in the mutant ida4. Six subspecies (all but f supported microtubule translocation with the maximal rate ranging from 2 to 12 μm s−1 and the apparent Km for ATP ranging from about 10 to 100 μM. All the subspecies translocated microtubules with the plus end leading, indicating that all the inner-arm dyneins are minus end-directed motors. Five subspecies (all but b and f) displayed microtubule rotation during translocation at rates of up to about 10 Hz. Unexpectedly, the Km values for ATP for translocation and rotation did not always agree; because of this, the pitch of the movement was variable with some subspecies. These observations indicate that axonemes are equipped with several inner-arm subspecies and that torque generation is a feature common to many of them.