Sex-specific binding and inactivation of agglutination factor in Chlamydomonas eugametos

Abstract

Gametes of opposite mating type (mt (+) and mt (-)) of the green alga Chlamydomonas eugametos agglutinate via their flagella as a prelude to sexual fusion. To quantitate sexual agglutination, an in vitro assay has been developed using (35)S-labeled flagella and the isolated mt (-)agglutination factor. It is shown that not only isolated flagella, but also the mt (-)agglutination factor rapidly bind to the flagella of intact gametes of the opposite mating type. This confirms the role of the mt (-)agglutination factor in determining the sexual agglutinability of mt (-)gametes. As a function of binding, the agglutinative power of the flagella of both mating types is destroyed by a temperature-sensitive process. Likewise, the mt (-)agglutination factor can be completely inactivated.

A monoclonal antibody that blocks adhesion of Chlamydomonas mt+ gametes.

“…Replacement of these lost adhesive sites requires protein synthesis (36, 37, 42) and protein glycosylation (29, 39, 48); Cooper et al showed that hydroxylation of proline was also required for replacement of these lost adhesion sites (10). Similar results have been shown for Chlamydomonas eugametos (32,33, 45,(47)(48)(49). To more fully study this adhesion-induced loss of flagellar adhesion sites and to identify molecules responsible for adhesion we have begun to use an immunological approach.…”

“…Recently Goodenough, Heuser, Adair, and co-workers identified high molecular mass glycoproteins rich in hydroxyproline that are involved in the adhesive interaction (1, 2, 4, 9, 10, 16). Saito and Matsuda (35, 37) obtained similar results using different purification methods, and van den Ende's laboratory has identified similar high molecular mass agglutinin molecules in C. eugametos (20,33, 45).Our laboratory has been interested in the molecular details of the adhesive interaction as well as a specific de-adhesion mechanism that we have characterized (39,(40)(41)(42)44). We showed that as cells adhere to each other they continuously inactivate the adhesiveness of each other's flagella (42, 44).…”

“…Recently Goodenough, Heuser, Adair, and co-workers identified high molecular mass glycoproteins rich in hydroxyproline that are involved in the adhesive interaction (1, 2, 4, 9, 10, 16). Saito and Matsuda (35, 37) obtained similar results using different purification methods, and van den Ende's laboratory has identified similar high molecular mass agglutinin molecules in C. eugametos (20,33, 45).…”

Cyclic AMP enhances the sexual agglutinability of Chlamydomonas flagella.

“…If, however, cell fusion cannot occur because of gene mutation or experimental manipulation, then adhesivity remains at elevated levels (5,10). During such protracted interactions, flagella are in fact continuously losing adhesivity (22,26), again by an unknown mechanism, and simultaneously regaining it, first from a preexisting pool of proteins and then by protein synthesis (4, 24,26). This loss/replacement cycle serves to maintain agglutinability at constant levels for many hours when cell fusion is inhibited.…”

Fluorophore-conjugated lectin labeling of the cell surface of isolated male and female gametes, central cells and synergids before and after fertilization in maize

“…In vitro fertilization experiments revealed that carbohydrates, as competing inhibitors, can prevent penetration of zona-free hamster and mouse eggs (Dravland and Mortimer 1988;Okabe et al 1989;Ponce et al 1994). In Chlamydomonas, lectins responsible for gamete adhesion have been identified and characterized on flagellar membranes (Musgrave et al 1981;Pijst et al 1984;Collin-Osdoby and Adair 1985;Klis et al 1985;Samson et al 1987). Wheat-germ agglutinin (WGA; Triticum vulgaris agglutinin), a lectin that binds to flagellar surfaces, induced all mating responses and increased intracellular cAMP in Chlamydomonas eugametos, suggesting an important function in gamete fusion and sexual signaling (Kooijman et al 1990).…”