Immunological comparison of 22S, 19S, and 12S dyneins from Paramecium cilia

Abstract

Three forms of dynein (22S, 19S, and 12S) were purified from Paramecium cilia. Two classes of monoclonal antibodies against purified 22S dynein were generated. One class reacted on immunoblots with the heavy chains of 22S, 19S, and 12S dyneins; the second class reacted with an 88 kD intermediate chain of 22S dynein. Polyclonal antiserum to the heavy chains of 22S dynein reacted with the alpha-heavy chain of 22S and 19S dyneins. A previously described antiserum raised against 22S dynein [Travis et al.: Biochim. Biophys. Acta 966:73-83, 1988] recognized the gamma-heavy chain of 22S dynein which was also present in 19S and 12S dyneins, along with the 88 and 76 kD intermediate chains of 22S dynein. This antiserum was also able to immunoprecipitate dynein from crude extracts of cilia. Electron microscopy revealed that the 22S dynein consisted mainly of two-headed particles with some three-headed particles present. The 12S dynein was mainly one-headed particles. The 19S dynein was a mixture of three-, two-, and one-headed particles. The immunological and electron microscopic studies showed that 19S dynein arises from 22S dynein, and that 12S dynein is heterogeneous, composed of the gamma-heavy chain of 22S dynein and a unique dynein ATPase. The polyclonal antibodies were also used to detect cross-reactive proteins in other organisms. Both the anti-heavy chain and the anti-22S dynein sera reacted strongly with 22S outer arm dynein of Tetrahymena, but not with the 14S dynein of this organism.