Cyclic AMP is involved in sexual reproduction of <i>Chlamydomonas eugametos</i>

Pijst, Hans L. A.; Driel, Roel van; Janssens, Pim M.W.; Musgrave, A.; Ende, H. van den

doi:10.1016/0014-5793(84)81091-1

Cited by 79 publications

(47 citation statements)

References 12 publications

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“…To do this we measured changes in cellular cAMP levels at various times during the mating reaction in the presence and absence of H-8 or staurosporine. Confirming earlier reports by Pijst et al (1984) and Pasquale and Goodenough (1987) there was a rapid, 8-10-fold increase of cellular cAMP within 2 rain after non-pretreated, control mt+ and mt-gametes were mixed together (Fig. 6, open circles).…”

Section: Staurosporine Inhibition Of Adhesion-induced Signaling In VIsupporting

confidence: 91%

“…During fertilization, cell-cell contacts between sperm and egg initiate G protein signaled responses in both cell types Gaffe, 1990;Kopf, 1990;Ward and Kopf, 1993). Although several molecules involved in cell-cell adhesion have been identified, the molecular mechanisms coupling adhe-adhesion is followed by a rapid increase of intracellular cAMP (Pijst et al, 1984;Pasquale and Goodenough, 1987;Kooijman et al, 1990) leading to several cellular responses required for continued adhesion as well as for preparation of the interacting cells for fusion and zygote formation (Goodenough, 1989;Snell et al, 1989;Kooijman et al, 1989;Tomson et al, 1990). This specific, cell-cell adhesion and signaling system is unique to gametes, and many of the signaled responses that occur in the cell body can be induced by incubation of gametes of a single mating type in dibutyryl cAMP (rib-CAMP) l (Pasquale and Goodenough, 1987;Snell et al, 1989;Goodenough, 1989;Hunnicutt et al, 1990).…”

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confidence: 99%

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Flagellar adhesion-dependent regulation of Chlamydomonas adenylyl cyclase in vitro: a possible role for protein kinases in sexual signaling.

Zhang

Snell

1994

The Journal of Cell Biology

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Abstract. Interactions between adhesion molecules, agglutinins, on the surfaces of the flagella of mt+ and mt-gametes in Ch/amydomonas rapidly generate a sexual signal, mediated by cAMP, that prepares the cells for fusion to form a zygote. The mechanism that couples agglutinin interactions to increased cellular levels of cAMP is unknown. In previous studies on the adenylyl cyclase in flagella of a single mating type (i.e., non-adhering flagella) we presented evidence that the gametic form of the enzyme, but not the vegetative form, was regulated by phosphorylation and dephosphorylation (Zhang, Y., E. M. Ross, and W. J. Snell. 1991. J. Biol. Chem. 266:22954-22959;Zhang, Y., and W. J. Snell. 1993. J. Biol. Chem. 268:1786-1791. In the present report we describe studies on regulation of flagellar adenylyl cyclase during adhesion in a cellfree system. The results show that the activity of gametic flagellar adenylyl cyclase is regulated by adhesion in vitro between flagella isolated from rat + and nat-gametes. After mixing rat + and rot-flagella together for 15 s in vitro, adenylyl cyclase activity was increased two-to threefold compared to that of the non-mixed (non-adhering), control flagella. This indicates that the regulation of gametic flagellar adenylyl cyclase during the early steps of fertilization is not mediated by signals from the cell body, but is a direct and primary response to interactions between mt ÷ and nat-agglutinins.By use of this in vitro assay, we discovered that 50 nM staurosporine (a protein kinase inhibitor) blocked adhesion-induced activation of adenylyl cyclase in vitro, while it had no effect on adenylyl cyelase activity of nowadhering gametic flagella. This same low concentration of staurosporine also inhibited adhesioninduced, increases in vivo in cellular cAMP and blocked subsequent cellular responses to adhesion. Taken together, our results indicate that flagellar adenylyl cyclase in Ch/amydomonas gametes is coupled to interactions between mt+ and rot-agglutinins by a staurosporine-sensitive activity, probably a protein kinase.THOUGH one result of an interaction between two developmentally primed cells can be the formation of a stable cell-to-cell adhesion, an equally important consequence often is the generation of a cellular signal. Cell contact-induced cellular signaling has been shown to be important during fertilization (Snell, 1990), in development of the nervous system (Kapfhammer and Schwab, 1992), in eye development in Drosophila (Hart et al., 1993), and in the immune system (Hynes, 1992;Dustin and Springer, 1991). During fertilization, cell-cell contacts between sperm and egg initiate G protein signaled responses in both cell types Gaffe, 1990;Kopf, 1990;Ward and Kopf, 1993). Although several molecules involved in cell-cell adhesion have been identified, the molecular mechanisms coupling adheAddress all correspondence to W. J. Snell, Department of Cell Biology and Neuroscience, University of Texas Southwestern Medical School, Dallas, TX 75235. sion to signal transduction are...

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Section: Staurosporine Inhibition Of Adhesion-induced Signaling In VIsupporting

confidence: 91%

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confidence: 99%

Flagellar adhesion-dependent regulation of Chlamydomonas adenylyl cyclase in vitro: a possible role for protein kinases in sexual signaling.

Zhang

Snell

1994

The Journal of Cell Biology

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“…To do this required a system in which flagellar adhesion did not interfere with our ability to examine mating structure interactions. We bypassed flagellar adhesion and focused directly on the interactions between mating structures by exploiting two features of the Chlamydomonas system: the availability of mutant mtϪ gametes (imp12) that do not express functional flagellar agglutinins (Pasquale and Goodenough, 1987;Goodenough, 1991;Goodenough et al, 1995a) and the ability to activate gametes of a single mating type experimentally by incubating them in dibutyryl cAMP (Pjist et al, 1984;Pasquale and Goodenough, 1987;Wilson et al, 1997). As shown in Figure 2A, flagellar adhesion between wild-type mtϩ and mtϪ gametes is characterized by close interactions between the flagella and accompanied by intimate contacts between the cell bodies at the sites of the activated mating structures.…”

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confidence: 99%

TheChlamydomonasFus1 Protein Is Present on the Mating TypeplusFusion Organelle and Required for a Critical Membrane Adhesion Event during Fusion withminusGametes

Misamore

Gupta

Snell³

2003

MBoC

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The molecular mechanisms of the defining event in fertilization, gamete fusion, remain poorly understood. The FUS1 gene in the unicellular, biflagellated green alga Chlamydomonas is one of the few sex-specific eukaryotic genes shown by genetic analysis to be essential for gamete fusion during fertilization. In Chlamydomonas, adhesion and fusion of the plasma membranes of activated mtϩ and mtϪ gametes is accomplished via specialized fusion organelles called mating structures. Herein, we identify the endogenous Fus1 protein, test the idea that Fus1 is at the site of fusion, and identify the step in fusion that requires Fus1. Our results show that Fus1 is a ϳ95-kDa protein present on the external surface of both unactivated and activated mtϩ gametes. Bioassays indicate that adhesion between mating type plus and mating type minus fusion organelles requires Fus1 and that Fus1 is functional only after gamete activation. Finally, immunofluorescence demonstrates that the Fus1 protein is present as an apical patch on unactivated gametes and redistributes during gamete activation over the entire surface of the microvillous-like activated plus mating structure, the fertilization tubule. Thus, Fus1 is present on mtϩ gametes at the site of cell-cell fusion and essential for an early step in the fusion process. INTRODUCTIONGamete fusion defines the beginning of a new organism in all sexual species. In most organisms, the events that precede fusion have been well characterized at the cellular level (Snell and White, 1996;Wassarman et al., 2001;Primakoff and Myles, 2002). Initial cell-cell interactions between sperm surface proteins and extracellular matrix molecules on the egg trigger activation of the sperm and exposure of previously cryptic regions of the plasma membrane proposed to be involved in gamete fusion. Once the sperm has made its way to the egg plasma membrane, the membranes of the two gametes interact more intimately, finally bringing about gamete fusion. Several sperm and egg molecules implicated in activation of the sperm have been identified, and we know much about the morphological events that accompany gamete fusion (Yanagimachi, 1994). On the other hand, little is known about the molecular mechanisms that underlie the late steps in fertilization, when the plasma membranes of the interacting gametes adhere and fuse (for review, see Primakoff and Myles, 2002). In the absence of any purely eukaryotic systems in which bona fide fusion proteins have been identified, the best models for fusion of the external plasma membrane of cells have come from studies of fusion of viruses with their eukaryotic target cells (White, 1996;Eckert and Kim, 2001). In these systems, a viral transmembrane surface protein interacts with a receptor in the host cell plasma membrane, leading to docking of the virus particle on the cell surface. Subsequent conformational changes in the interacting proteins finally lead to complete fusion (Dimitrov, 2000;Malashkevich et al., 2001;Mayer, 2001).A small number of eukaryotic genes has been sho...

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“…In addition to binding the flagella of gametes of opposite mating types to each other, these receptor/ligand-like interactions also initiate a complex series of events termed gamete activation. One of the earliest documented steps in gamete activation triggered by agglutinin interactions is activation of a gamete-specific flagellar adenylyl cyclase and generation of cAMP (7)(8)(9)(10)(11). As part of an intricate feedback mechanism required to maintain and enhance flagellar adhesiveness and to keep the cells bound to each other until cell-cell fusion occurs, cAMP levels increase in the cell bodies of the gametes of both mating types during flagellar adhesion and lead to increased flagellar adhesiveness (6,9,12,13).…”

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Regulated Targeting of a Protein Kinase into an Intact Flagellum

Pan

Snell

2000

Journal of Biological Chemistry

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In the green alga Chlamydomonas reinhardtii flagellar adhesion between gametes of opposite mating types leads to rapid cellular changes, events collectively termed gamete activation, that prepare the gametes for cell-cell fusion. As is true for gametes of most organisms, the cellular and molecular mechanisms that underlie gamete activation are poorly understood. Here we report on the regulated movement of a newly identified protein kinase, Chlamydomonas aurora/Ipl1p-like protein kinase (CALK), from the cell body to the flagella during gamete activation. CALK encodes a protein of 769 amino acids and is the newest member of the aurora/ Ipl1p protein kinase family. Immunoblotting with an anti-CALK antibody showed that CALK was present as a 78/80-kDa doublet in vegetative cells and unactivated gametes of both mating types and was localized primarily in cell bodies. In cells undergoing fertilization, the 78-kDa CALK was rapidly targeted to the flagella, and within 5 min after mixing gametes of opposite mating types, the level of CALK in the flagella began to approach levels normally found in the cell body. Protein synthesis was not required for targeting, indicating that the translocated CALK and the cellular molecules required for its movement are present in unactivated gametes. CALK was also translocated to the flagella during flagellar adhesion of nonfusing mutant gametes, demonstrating that cell fusion was not required for movement. Finally, the requirement for flagellar adhesion could be bypassed; incubation of cells of a single mating type in dibutyryl cAMP led to CALK translocation to flagella in gametes but not vegetative cells. These experiments document a new event in gamete activation in Chlamydomonas and reveal the existence of a mechanism for regulated translocation of molecules into an intact flagellum.Cell-cell interactions leading to fusion between gametes of opposite sexes during fertilization are complex processes that involve dramatic changes in each of the interacting gametes. In most multicellular organisms, interactions between adhesion molecules/receptors on the sperm plasma membrane and ligands on the egg surface activate poorly understood signaling pathways that bring about transformation of the sperm into a fusion competent, activated gamete (1). For example, the sperm surface is remodeled as a consequence of the acrosome reaction, an event that accompanies gamete activation, and previously existing adhesion molecules are mobilized to new sites on the sperm (2-4). The molecular mechanisms that underlie and regulate signal transduction and movement of molecules between different gamete compartments during gamete activation largely are unknown (2).As in multicellular organisms, gamete activation and fertilization in the unicellular green alga Chlamydomonas (5) depend on adhesion-induced signaling pathways and intercompartmental communication. At the completion of gametogenesis, during which asexually growing mtϩ and mtϪ vegetative cells differentiate into sexually competent cells, the resultin...

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Cyclic AMP is involved in sexual reproduction of Chlamydomonas eugametos

Cited by 79 publications

References 12 publications

Flagellar adhesion-dependent regulation of Chlamydomonas adenylyl cyclase in vitro: a possible role for protein kinases in sexual signaling.

Flagellar adhesion-dependent regulation of Chlamydomonas adenylyl cyclase in vitro: a possible role for protein kinases in sexual signaling.

TheChlamydomonasFus1 Protein Is Present on the Mating TypeplusFusion Organelle and Required for a Critical Membrane Adhesion Event during Fusion withminusGametes

Regulated Targeting of a Protein Kinase into an Intact Flagellum

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