Biochemical studies of the excitable membrane of Paramecium tetraurelia. III. Proteins of cilia and ciliary membranes.

Abstract

The protein composition of the ciliary membrane of wild-type cells was defined. Membranes of high purity and integrity were obtained without the use of detergents. The fractions were characterized by EM and the proteins of whole cilia, axonemes and ciliary membrane vesicles were resolved by SDS [sodium dodecylsulfate] polyacrylamide gel electrophoresis and isoelectric focusing in 1 and 2 dimensions. Protein patterns and EM appearance of the fractions were highly reproducible. Over 200 polypeptides were present in isolated cilia, most of which were recovered in the axonemal fraction. Trichocysts, which were sometimes present as a minor contaminant in ciliary preparations, were composed of a very distinct set of over 30 polypeptides of MW 11,000-19,000. Membrane vesicles contained up to 70 polypeptides of MW 15,000-250,000. The major vesicle species were a high MW protein (the immobilization antigen) and a group of acidic proteins with MW .apprx. 40,000. These and several other membrane proteins were specifically decreased or totally absent in the axoneme fraction. Tubulin, the major axonemal species, occurred only in trace amounts in isolated vesicles; the same was true for Tetrahymena ciliary membranes prepared by the methods described in this paper. A protein of MW 31,000, pI 6.8, was virtually absent in vesicles prepared from cells in exponential growth phase, but became prominent early in stationary phase in good correlation with cellular mating reactivity. This detailed characterization will provide the basis for comparison of the ciliary proteins of wild-type and behavioral mutants and for analysis of topography and function of membrane proteins. It will also be useful in future studies of trichocysts and mating reactions.