Ciliary polypeptides and glycoconjugates of wild‐type and mutant Tetrahymena thermophila: Starved versus nonstarved

Abstract

To investigate the role of cilia in mating interactions of Tetrahymena thermophila, ciliary membrane-rich fractions were isolated from two wild-type strains, a non-discharge mucocyst mutant which possesses mating behavior similar to wild-type, and a mating mutant which is able to costimulate cells of complementary mating type but cannot enter into pair formation. In each case, proteins from the ciliary membrane-rich fractions of starved, mating-competent ("initiated") cells were compared with those from non-starved, mating-incompetent ("non-initiated") cells, by gel electrophoresis and lectin blotting. In stained gels, a 43 kDa polypeptide was reduced or absent in initiated cells but present in non-initiated cells, in all strains. In silver-stained gels, a 25 kDa polypeptide was present in all strains, both initiated and non-initiated. In blots probed with Con A-peroxidase, a 25 kDa glycoprotein was present in ciliary membrane fractions from non-initiated cells and absent in membranes of initiated cells of the two wild-type strains and the mucocyst mutant, but is present in initiated and non-initiated cells of the mating mutant (several hypotheses are presented to explain these findings). In addition, ciliary proteins of the mating mutant included at least two unique Con A-binding polypeptides. Our results support the idea that development of mating competence during starvation involves an extensive remodeling of ciliary membranes, and identify a 25 kDa glycoconjugate as having a potential role in control of pair formation during mating.

Affinity‐Purification of Concanavalin A‐Binding Ciliary Glycoconjugates of Starved and Feeding Tetrahymena thermophila

“…General effects of starvation on extractable polypeptides and glycoconjugates. Our results, based on different methods for preparation and analysis of Con A-binding ciliary membrane glycoproteins of WT T. thermophila, provide independent evidence corroborating our earlier published and unpublished findings [6]. We confirm that during starvation there is an overall change in the pattern of Con A-binding glycoproteins, affecting many but not all proteins of WT cells (see Figs.…”

“…3). Cheng and Hufnagel [6] also found differences between WT and mutant cells, and reported two prominent bands at 66 kDa and 94 kDa in mutant but not WT cells. These differences may explain why the mutant is unable to enter into pair formation.…”

“…There is also direct evidence, through inhibitor and labeling studies, as well as biochemical analysis, that Con A-binding sites are present on the surface of the cilia and that these binding sites may be important in mating [5, 7, To identify Con A-binding ciliary membrane proteins that might be involved in early recognition events of mating in T. thermophila, we initiated a study of ciliary membrane glycoproteins and their responses to starvation, using mating-competent wild type cells and a mating mutant unable to enter into pair formation. Using lectin blotting, we previously found evidence for the presence of a 25 kDa glycoprotein with a potential regulatory role in the development of mating reactivity [6] (Hufnagel, L. A. 1993.…”

Comparative Genomic Analysis of Integral Membrane Transport Proteins in Ciliates

“…; Wereszka and Michalowski ). The paucity of polysaccharide export proteins could be explained if ciliates use exocytosis for polysaccharide export (Cheng and Hufnagel ; Gutierrez and Orias ; Turkewitz ). The studies reported here provide a wealth of novel information relevant to the ecology, evolution, physiology, and pathology of model ciliates and probably to the characteristics of many other ciliates of industrial and bioremediative importance (Dubber and Gray ; Gertler et al.…”

Evolution of intraflagellar transport from coated vesicles and autogenous origin of the eukaryotic cilium

“…(61,62) Agglutination during mating is also mediated by cilia in ciliates (63) (Paramecium, Tetrahymena, Euplotes) with a potential role of ciliary glycoproteins in pair formation. (64,65) Ca 2þ -and cAMPdependent signalling is also activated during sexual agglutination in Chlamydomonas. To decide whether ciliary attachment is ancestrally associated with cilia, the role of cilia in the mating of ciliated amoebozoa and choanoflagellates should be studied.…”