Aggregation-dependent turnover of flagellar adhesion molecules in Chlamydomonas gametes

Previous studies on flagellar adhesion in Chlamydomonas (Snell, W. and S. Roseman. 1979. J. Biol. Chem. 254: 10820-10829.) have shown that as gametes adhere to flagella isolated from gametes of the opposite mating type, the adhesiveness of the added flagella but not of the gametes is lost. The studies reported here show that the addition of protein synthesis inhibitors (cycloheximide [CH] or anisomycin) to the medium of such cell-flagella mixtures causes the cells to lose their adhesiveness. This loss, however, occurs only after the cells have interacted with 4-8 flagella/cell and does not occur if the cells are kept in CH (7 h) without aggregating. The availability of an impotent (imp) mating type plus (mt⁺) mutant (provided by U. W. Goodenough), which adheres but is unable to undergo the fusion that normally follows adhesion, made it possible to determine whether a similar loss of adhesiveness occurs in mixtures of mating type minus (mt⁻) and imp mt⁺ gametes. In the absence of inhibitor, mt⁻ and imp mt⁺ gametes adhered to each other (without fusing) for several hours; however, in the presence of CH or anisomycin, the gametes began to de-adhere 35 min after mixing, and, by 90 min, 100 of the cells were single again. This effect was reversible, and the rapid turnover of molecules involved in adhesion ocurred only during adhesion inasmuch as gametes pretreated for 4 h with CH were able to aggregate in CH for the same length of time as nonpretreated cells aggregated in CH. By the addition of CH at various times after the mt⁻ and imp mt⁺ gametes were mixed, measurements were made of the "pool size" of the molecules involved in adhesion. The pool reached a minimum after 25 min of aggregation, rapidly increased for the next 25 min, and then leveled off at the premixing level. These results suggest that flagellar adhesion in Chlamydomonas causes modification of surface molecules (receptors, ligands), which brings about their inactivation and stimulates their replacement.