Feedback inhibition of Ras activity coordinates cell fusion with cell-cell contact

AbstractIn fission yeast Schizosaccharomyces pombe, pheromone signalling engages a GPCR-Ras-MAPK cascade to trigger sexual differentiation leading to gamete fusion. Cell-cell fusion necessitates local cell wall digestion, the location of which relies on an initially dynamic actin fusion focus that becomes stabilized upon local enrichment of the signalling cascade. We constructed a live-reporter of active Ras1 (Ras1-GTP), also functional in S. cerevisiae, which revealed Ras activity at polarity sites peaking on the fusion structure before fusion. Remarkably, constitutive Ras1 activation promoted fusion focus stabilization and fusion attempts irrespective of cell-cell pairing, leading to cell lysis. Ras1 activity is restricted by the GTPase activating protein (GAP) Gap1, itself recruited to sites of Ras1-GTP. While the GAP domain on its own does not suffice for this localization, its recruitment to Ras1-GTP sites is essential to block untimely fusion attempts. We conclude that negative feedback control of Ras activity restrains the MAPK signal and couples fusion with cell-cell engagement.

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Inhibition of Ras activity coordinates cell fusion with cell–cell contact during yeast mating

The Journal of Cell Biology ◽

10.1083/jcb.201708195 ◽

2018 ◽

Vol 217 (4) ◽

pp. 1467-1483 ◽

Cited By ~ 18

Author(s):

Laura Merlini ◽

Bita Khalili ◽

Omaya Dudin ◽

Laetitia Michon ◽

Vincent Vincenzetti ◽

...

Keyword(s):

Cell Fusion ◽

Mitogen Activated Protein Kinase ◽

Cell Contact ◽

Pheromone Signaling ◽

Mitogen Activated Protein ◽

Fusion Cell ◽

Local Cell ◽

Guanosine Triphosphatase ◽

Local Enrichment ◽

Cell Cell

In the fission yeast Schizosaccharomyces pombe, pheromone signaling engages a signaling pathway composed of a G protein–coupled receptor, Ras, and a mitogen-activated protein kinase (MAPK) cascade that triggers sexual differentiation and gamete fusion. Cell–cell fusion requires local cell wall digestion, which relies on an initially dynamic actin fusion focus that becomes stabilized upon local enrichment of the signaling cascade on the structure. We constructed a live-reporter of active Ras1 (Ras1–guanosine triphosphate [GTP]) that shows Ras activity at polarity sites peaking on the fusion structure before fusion. Remarkably, constitutive Ras1 activation promoted fusion focus stabilization and fusion attempts irrespective of cell pairing, leading to cell lysis. Ras1 activity was restricted by the guanosine triphosphatase–activating protein Gap1, which was itself recruited to sites of Ras1-GTP and was essential to block untimely fusion attempts. We propose that negative feedback control of Ras activity restrains the MAPK signal and couples fusion with cell–cell engagement.

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Studies of membrane fusion. I. Paramyxovirus-induced cell fusion, a scanning electron-microscope study

Journal of Cell Science ◽

10.1242/jcs.28.1.179 ◽

1977 ◽

Vol 28 (1) ◽

pp. 179-188

Author(s):

S. Knutton ◽

D. Jackson ◽

M. Ford

Keyword(s):

Cell Surface ◽

Cell Fusion ◽

Hela Cells ◽

Cell Swelling ◽

Cell Contact ◽

Cell Agglutination ◽

Scanning Electron Microscope Study ◽

Virus Particles ◽

Scanning Electron ◽

Cell Cell

Fusion of erythrocytes and HeLa cells with Sendai and Newcastle disease viruses has been studied by scanning electron microscopy. Most virus particles are spherical but vary in diameter from approximately 200 to approximately 600 nm. At 4 degrees C virus particles bind randomly to the cell surface and at high cell densities cross-linking of adjacent cells by virus particles results in cell agglutination. Cell-cell fusion takes place when the agglutinated cell suspension is warmed to 37 degrees C. Fusion is initiated at sites of cell-cell contact and is accompanied in all cases by cell swelling. In the case of suspension HeLa cells, virally mediated cell swelling involves an ‘unfolding’ of cell surface microvilli and results in the formation of smooth-surfaced single or fused cells. With erythrocytes, swelling results in haemolysis. There is a dramatic reduction in the numbers of virus particles bound to cells following fusion.

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Osmotic Balance Regulates Cell Fusion during Mating in Saccharomyces cerevisiae

The Journal of Cell Biology ◽

10.1083/jcb.138.5.961 ◽

1997 ◽

Vol 138 (5) ◽

pp. 961-974 ◽

Cited By ~ 102

Author(s):

Jennifer Philips ◽

Ira Herskowitz

Keyword(s):

Cell Wall ◽

Cell Fusion ◽

Cell Contact ◽

Fusion Event ◽

Glycerol Facilitator ◽

Fusion Defect ◽

Osmotic Balance ◽

Glycerol 3 Phosphate Dehydrogenase ◽

Gpd1 Gene ◽

Cell Cell

Successful zygote formation during yeast mating requires cell fusion of the two haploid mating partners. To ensure that cells do not lyse as they remodel their cell wall, the fusion event is both temporally and spatially regulated: the cell wall is degraded only after cell–cell contact and only in the region of cell–cell contact. To understand how cell fusion is regulated, we identified mutants defective in cell fusion based upon their defect in mating to a fus1 fus2 strain (Chenevert, J., N. Valtz, and I. Herskowitz. 1994. Genetics 136:1287–1297). Two of these cell fusion mutants are defective in the FPS1 gene, which codes for a glycerol facilitator (Luyten, K., J. Albertyn, W.F. Skibbe, B.A. Prior, J. Ramos, J.M. Thevelein, and S. Hohmann. 1995. EMBO [Eur. Mol. Biol. Organ.] J. 14:1360–1371). To determine whether inability to maintain osmotic balance accounts for the defect in cell fusion in these mutants, we analyzed the behavior of an fps1Δ mutant with reduced intracellular glycerol levels because of a defect in the glycerol-3-phosphate dehydrogenase (GPD1) gene (Albertyn, J., S. Hohmann, J.M. Thevelein, and B.A. Prior. 1994. Mol. Cell. Biol. 14:4135– 4144): deletion of GPD1 partially suppressed the cell fusion defect of fps1 mutants. In contrast, overexpression of GPD1 exacerbated the defect. The fusion defect could also be partially suppressed by 1 M sorbitol. These observations indicate that the fusion defect of fps1 mutants results from inability to regulate osmotic balance and provide evidence that the osmotic state of the cell can regulate fusion. We have also observed that mutants expressing hyperactive protein kinase C exhibit a cell fusion defect similar to that of fps1 mutants. We propose that Pkc1p regulates cell fusion in response to osmotic disequilibrium. Unlike fps1 mutants, fus1 and fus2 mutants are not influenced by expression of GPD1 or by 1 M sorbitol. Their fusion defect is thus unlikely to result from altered osmotic balance.

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