Identification and visualization of the sexual agglutinin from the mating-type plus flagellar membrane of Chlamydomonas

Adair, W S; Hwang, Carol; Goodenough, Ursula

doi:10.1016/0092-8674(83)90347-1

Cited by 89 publications

(42 citation statements)

References 23 publications

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“…We used seven Chlamydomonas reinhardtii strains in this study: wild-type strains CC-620 (mt1) and CC-124 (mt2), the agglutinin 1 defective mutant imp-5 (CC-469; Adair et al, 1983), a mutant deficient in the accumulation of cAMP (imp-3, CC-465;Goodenough et al, 1976), the fusion-defective mutant fus1 (CC-1158; Ferris et al, 1996), and the flagellarless mutant bld2 (Goodenough and St. Clair, 1975), all obtained from the Chlamydomonas Genetics Center (Duke University, Durham, NC). The fusion-defective mutant ida5 (mt1) was kindly provided by R. Kamiya (Kato-Minoura et al, 1997).…”

Section: Strains and Cell Culturementioning

confidence: 99%

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Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas

Hoffmann

Beck

2005

Plant Physiology

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The first step in sexual differentiation of the unicellular green alga Chlamydomonas reinhardtii is the formation of gametes. Three genes, GAS28, GAS30, and GAS31, encoding Hyp-rich glycoproteins that presumably are cell wall constituents, are expressed in the late phase of gametogenesis. These genes, in addition, are activated by zygote formation and cell wall removal and by the application of osmotic stress. The induction by zygote formation could be traced to cell wall shedding prior to gamete fusion since it was seen in mutants defective in cell fusion. However, it was absent in mutants defective in the initial steps of mating, i.e. in flagellar agglutination and in accumulation of adenosine 3#,5#-cyclic monophosphate in response to this agglutination. Induction of the three GAS genes was also observed when cultures were exposed to hypoosmotic or hyperosmotic stress. To address the question whether the induction seen upon cell wall removal from both gametes and vegetative cells was elicited by osmotic stress, cell wall removal was performed under isosmotic conditions. Also under such conditions an activation of the genes was observed, suggesting that the signaling pathway(s) is (are) activated by wall removal itself.

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Section: Strains and Cell Culturementioning

confidence: 99%

“…The initial contact between gametes of mating types plus and minus is mediated by the mating-typespecific agglutinins, high M r glycoproteins on the flagellar surface (Adair et al, 1983;Ferris et al, 2005). The resulting flagellar adhesion generates signaling pathways that activate the interacting cells for cell-cell fusion.…”

mentioning

confidence: 99%

Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas

Hoffmann

Beck

2005

Plant Physiology

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“…The OG extracts were fractionated by molecular sieve chromatography using a Biogel A-15M column as described in Materials and Methods. Adhesive activity was separated from mastigoneme protein and the major membrane protein with an Mr of ~350,000, which has been shown by us and others (2,6,16,19,38,50) to be the major protein component of flagellar surface fractions (not shown). Typically, this purification method yielded 60-100% of the starting activity with a 20-30-fold increase in specific activity.…”

Section: Partial Purification Of Adhesion Molecules From Mt + Gametesmentioning

confidence: 99%

“…Several laboratories have been interested in the initial adhesive interaction. 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).…”

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confidence: 99%

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

Snell¹,

Clausell

Perillo³

et al. 1986

The Journal of Cell Biology

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Abstract. During the mating reaction (fertilization) in the biflagellated alga, Chlamydomonas reinhardtii, mt* and mt-gametes adhere to each other via their flagella and subsequently fuse to form quadriflagellated zygotes. In the studies reported here, we describe a monoclonal antibody directed against an mt + flagellar surface molecule. The antibody blocks the adhesiveness of mt + gametes, isolated mt ÷ flagella, and detergent extracts thereof. It has no effect on mt-gametes. Cyanogen bromide-activated Sepharose beads derivatized with the antibody bind only mt + gametes; mt-gametes and mt ÷ and mt-vegetative cells are unreactive with the derivatized beads. The interaction of mt + gametes with the beads is dynamic and cells continuously bind, detach, and rebind to the beads. Surprisingly, antibody-derivatized beads that have been incubated with mt + gametes acquire the ability to bind mr-gametes. Moreover, extraction of the preincubated beads with detergents releases active mt + adhesion molecules. The evidence suggests that binding of the antibody to the flagellar surface adhesion molecules causes their release from the flagellar surface, possibly mimicking the normal mechanism of flagellar de-adhesion.W HEN gametes of opposite mating types of the biflagellated alga Chlamydomonas reinhardtii are mixed together, they adhere to each other via their flagella and form large clumps of aggregating cells. Within these clumps, pairs of cells form that eventually fuse to become quadriflagellated zygotes. Several laboratories have been interested in the initial adhesive interaction. 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). 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. In earlier unpublished studies we had attempted to prepare rabbit polyclonal antibodies against flagella or flagellar surface fractions that would specifically block adhesiveness of mating type plus (mr+) 1 or mating type minus (mr-) gametes. To date, all of th...

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“…The plus and minus versions from C. reinhardtii are encoded by different genes and possess complementary adhesive properties, but they also share common features: (1) they are both huge monomeric glycoproteins (native molecular mass > 1000 kD; Adair et al, 1983); (2) their association with the flagellar surface is disrupted by EDTA (Adair et al, 1982), indicating that they are extrinsic membrane proteins; (3) they are both chimeric (Kieliszewski and Lamport, 1994), possessing a large globular head and a fibrous shaft ; (4) they are members of the Hyprich glycoprotein (HRGP) family (Cooper et al, 1983) found as well in the cell walls of most green organisms (reviewed in Cassab, 1998;Serpe and Nothnagel, 1999) and implicated in sexual interactions in higher plants (reviewed in Wu et al, 2001).…”

Section: Introductionmentioning

confidence: 99%

PlusandMinusSexual Agglutinins fromChlamydomonas reinhardtii

et al. 2005

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Gametes of the unicellular green alga Chlamydomonas reinhardtii undergo sexual adhesion via enormous chimeric Hyp-rich glycoproteins (HRGPs), the plus and minus sexual agglutinins, that are displayed on their flagellar membrane surfaces. We have previously purified the agglutinins and analyzed their structural organization using electron microscopy. We report here the cloning and sequencing of the Sag1 and Sad1 genes that encode the two agglutinins and relate their derived amino acid sequences and predicted secondary structure to the morphology of the purified proteins. Both agglutinin proteins are organized into three distinct domains: a head, a shaft in a polyproline II configuration, and an N-terminal domain. The plus and minus heads are related in overall organization but poorly conserved in sequence except for two regions of predicted hydrophobic a-helix. The shafts contain numerous repeats of the PPSPX motif previously identified in Gp1, a cell wall HRGP. We propose that the head domains engage in autolectin associations with the distal termini of their own shafts and suggest ways that adhesion may involve head-head interactions, exolectin interactions between the heads and shafts of opposite type, and antiparallel shaft-shaft interactions mediated by carbohydrates displayed in polyproline II configurations.

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Identification and visualization of the sexual agglutinin from the mating-type plus flagellar membrane of Chlamydomonas

Cited by 89 publications

References 23 publications

Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas

Mating-Induced Shedding of Cell Walls, Removal of Walls from Vegetative Cells, and Osmotic Stress Induce Presumed Cell Wall Genes in Chlamydomonas

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

PlusandMinusSexual Agglutinins fromChlamydomonas reinhardtii

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