Gametic differentiation in Chlamydomonas reinhardtii. II. Flagellar membranes and the agglutination reaction.

Abstract

A structural and biochemical study is presented concerning the agglutination of gametic flagella, the initial step in the mating reaction of Chlamydomonas reinhardtii. An alteration in the distribution of the intramembranous particles revealed by freeze-fracturing of flagellar membranes is shown to accompany gametic differentiation in both mating types. The isolation and electrophoretic analysis of flagellar membranes and mastigonemes are reported; no electrophoretic differences can be detected when the membrane or mastigoneme glycoproteins from vegetative and gametic cells are compared, nor when glycoproteins from the two mating types are compared, and no novel polypeptides are present in gametic preparations. The membrane vesicles, after they are freed of mastigonemes by sedimentation through a discontinuous sucrose gradient, are extremely active as an isoagglutinin, indicating a direct involvement of the membrane in the mating reaction.As cells differentiate, they are often conferred with unique surface properties that allow extremely selective cellular interactions: the tissue-specific adhesion of dissociated embryonic cells (1), the species-specific reaggregation of dissociated sponge cells (16), and the developmental cycle of agglutinability in Dictyostelium (3) are three prominent examples. Such selective interactions have also been observed in or inferred from studies of simple eukaryotic organisms that undergo mating reactions. Thus, the composition or architecture of the cell surface appears to confer sexual discrimination in Hansenula wingii (7), in Paramecium (29), and in many of the volvocid phytoflagellates which agglutinate through the contact of their flagellar surfaces (31).In this paper we present the results of our observations on the flagellar surface of the phytoflagellate Chlam.vdomonas reinhardtii, which develops the capacity for such specific agglutination reactions after a simple alteration of its growth medium. Specifically, when vegetative C, reinhardtii cells of opposite mating types (rot + and rot-) are induced to differentiate into gametes by nitrogen starvation (19,25) and are mixed together, they agglutinate by their flagellar tips as a prelude to the zygotic cell fusion that occurs several minutes later (described in reference 13). The flagella of vegetative (nongametic) cells exhibit no such agglutinability, nor do certain nonmating mutant strains that have been isolated in this laboratory. Therefore, it should be possible to identify the agglutinins active in this reaction and to study both their mode of action and the 606

PlusandMinusSexual Agglutinins fromChlamydomonas reinhardtii

“…In pioneering studies, Wiese documented that the mating-typespecific components that induce mating-type plus gametic flagella to adhere to mating-type minus gametic flagella could be recovered from the culture medium (Fö rster and Wiese, 1954;Wiese, 1965). Subsequent analyses of such gamone preparations revealed them to contain flagellar membrane vesicles (Bergman et al, 1975;Snell, 1976;Musgrave et al, 1981), and further studies eventually led to the purification and extensive characterization of their component agglutinin proteins in both C. reinhardtii and the distantly related C. eugametos Cooper et al, 1983;Collin-Osdoby et al, 1984;Adair, 1985;Collin-Osdoby and Adair, 1985;Samson et al, 1987;Homan et al, 1988;Goodenough and Adair, 1989).…”

Gametic differentiation in Chlamydomonas reinhardtii. I. Production of gametes and their fine structure.

“…We conclude that many of these changes are probably degradative events stimulated by nitrogen starvation and/or events geared to insure the long-term survival of gametes in the face of a nitrogen-free environment, and we define those few phenotypes that are directly related to the acquisition of mating ability. Two of these gamete-specific traits are explored more extensively in the accompanying papers in this series (1,9).…”

Functional modification of the Chlamydomonas flagellar surface.

“…The flagellar surface consists of an extension of the cell body plasma membrane which is covered by a 20-nm layer of material referred to as the "flagellar sheath" (27) or "fuzzy coat" (2) in addition to mastigo-88 ROBERT A. BLOODGOOD and GREGORY S. MAY Department of Anatomy, University of Virginia School of Medicine, Charlottesville, Virginia 22908, and the Department of Biology, Yale University, New Haven, Connecticut 06520 nemes -0.91ím in length (27,35). Purified flagellar membranes consist of one predominant class of high molecular weight (HMW) glycoproteins in addition to a number of minor components (2,19,22,23,29,35) .Chlamydomonas can be grown vegetatively or induced to differentiate into gametic cells. Vegetative and gametic cells possess certain flagellar surface properties in common although flagella from gametic cells express certain unique properties (6, 11) .…”