Properties of microtubule sliding disintegration in isolated tetrahymena cilia

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Abstract
Properties of the sliding disintegration response of demembranated T. pyriformis cilia were studied by measuring the spectrophotometric response or turbidity of cilia suspensions at a wavelength of 350 nm relative to changes in the dynein substrate (MgATP2-) concentration. The maximum decrease in turbidity occurs in 20 .mu.M ATP, and 90% of the decrease occurs in .apprx. 5.9 s. At lower ATP concentrations (1-20 .mu.M), both the velocity and magnitude of the turbidity decreases are proportional to ATP concentration. The velocity data for 20 .mu.M ATP permit construction of a reaction velocity curve, suggesting that changes in turbidity are directly proportional to the extent and velocity of disintegration. At ATP concentration > 20 .mu.M (50 .mu.M to 5 mM), both velocity and magnitude of the turbidimetric response are reduced by .apprx. 50%. This apparent inhibition results in a biphasic response curve that may be related to activation of residual shear resistance or regulatory components at the higher ATP concentrations. The inhibitory effects of elevated ATP can be eliminated by mild trypsin proteolysis, where the reaction goes to completion at any ATP concentration. The turbidimetric responses of the axoneme-substrate suspensions are consistent with the extent and type of axoneme disintegration revealed by EM examination of the various suspensions, suggesting that the turbidimetric assay may be a reliable means for assessing the state of axoneme integrity.