scholarly journals Ultrastructural plasma membrane asymmetries in tension and curvature promote yeast cell fusion

2021 ◽  
Vol 220 (10) ◽  
Author(s):  
Olivia Muriel ◽  
Laetitia Michon ◽  
Wanda Kukulski ◽  
Sophie G. Martin
Keyword(s):  
Plasma Membrane ◽  
Cell Fusion ◽  
Turgor Pressure ◽  
Light And Electron Microscopy ◽  
Asymmetric Membrane ◽  
Local Reduction ◽  
Fusion Site ◽  
Actin Structure ◽  
Fusion Pores ◽  
Cell Cell

Cell–cell fusion is central for sexual reproduction, and generally involves gametes of different shapes and sizes. In walled fission yeast Schizosaccharomyces pombe, the fusion of h+ and h− isogametes requires the fusion focus, an actin structure that concentrates glucanase-containing vesicles for cell wall digestion. Here, we present a quantitative correlative light and electron microscopy (CLEM) tomographic dataset of the fusion site, which reveals the fusion focus ultrastructure. Unexpectedly, gametes show marked asymmetries: a taut, convex plasma membrane of h− cells progressively protrudes into a more slack, wavy plasma membrane of h+ cells. Asymmetries are relaxed upon fusion, with observations of ramified fusion pores. h+ cells have a higher exo-/endocytosis ratio than h− cells, and local reduction in exocytosis strongly diminishes membrane waviness. Reciprocally, turgor pressure reduction specifically in h− cells impedes their protrusions into h+ cells and delays cell fusion. We hypothesize that asymmetric membrane conformations, due to differential turgor pressure and exocytosis/endocytosis ratios between mating types, favor cell–cell fusion.

scholarly journals Ultrastructural plasma membrane asymmetries in tension and curvature promote yeast cell fusion

2021 ◽  
Author(s):  
Olivia Muriel ◽  
Laetitia Michon ◽  
Wanda Kukulski ◽  
Sophie G Martin
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Sexual Reproduction ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Turgor Pressure ◽  
Wall Material ◽  
M Cells ◽  
P Cells ◽  
Cell Cell

Cell-cell fusion is central to the process of fertilization for sexual reproduction. This necessitates the remodeling of peri-cellular matrix or cell wall material and the merging of plasma membranes. In walled fission yeast S. pombe, the fusion of P and M cells during sexual reproduction relies on the fusion focus, an actin structure that concentrates glucanase-containing secretory vesicles for local cell wall digestion necessary for membrane fusion. Here, we present a correlative light and electron microscopy (CLEM) quantitative study of a large dataset of 3D tomograms of the fusion site, which revealed the ultrastructure of the fusion focus as an actin-containing, vesicle-dense structure excluding other organelles. Unexpectedly, the data revealed asymmetries between the two gametes: M-cells exhibit a taut and convex plasma membrane that progressively protrudes into P-cells, which exhibit a more slack, wavy plasma membrane. These asymmetries are relaxed upon plasma membrane fusion, with observations of ramified pores that may result from multiple initiations or inhomogeneous expansion. We show that P-cells have a higher exo- to endocytosis ratio than M-cells, and that local reduction in exocytosis abrogates membrane waviness and compromises cell fusion significantly more in P- than M-cells. Reciprocally, reduction of turgor pressure specifically in M-cells prevents their protrusions into P-cells and delays cell fusion. Thus, asymmetric membrane conformations, which result from differential turgor pressure and exocytosis/endocytosis ratios between mating types, favor cell-cell fusion.

scholarly journals FUS1Regulates the Opening and Expansion of Fusion Pores between Mating Yeast

2006 ◽  
Vol 17 (5) ◽  
pp. 2439-2450 ◽  
Author(s):  
Scott Nolan ◽  
Ann E. Cowan ◽  
Dennis E. Koppel ◽  
Hui Jin ◽  
Eric Grote
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Yeast Cells ◽  
Fusion Pore ◽  
Sudden Increase ◽  
Fluorescent Markers ◽  
Fold Reduction ◽  
Fusion Pores

Mating yeast cells provide a genetically accessible system for the study of cell fusion. The dynamics of fusion pores between yeast cells were analyzed by following the exchange of fluorescent markers between fusion partners. Upon plasma membrane fusion, cytoplasmic GFP and DsRed diffuse between cells at rates proportional to the size of the fusion pore. GFP permeance measurements reveal that a typical fusion pore opens with a burst and then gradually expands. In some mating pairs, a sudden increase in GFP permeance was found, consistent with the opening of a second pore. In contrast, other fusion pores closed after permitting a limited amount of cytoplasmic exchange. Deletion of FUS1 from both mating partners caused a >10-fold reduction in the initial permeance and expansion rate of the fusion pore. Although fus1 mating pairs also have a defect in degrading the cell wall that separates mating partners before plasma membrane fusion, other cell fusion mutants with cell wall remodeling defects had more modest effects on fusion pore permeance. Karyogamy is delayed by >1 h in fus1 mating pairs, possibly as a consequence of retarded fusion pore expansion.

scholarly journals Evolutionarily related small viral fusogens hijack distinct but modular actin nucleation pathways to drive cell-cell fusion

2020 ◽  
Vol 118 (1) ◽  
pp. e2007526118
Author(s):  
Ka Man Carmen Chan ◽  
Ashley L. Arthur ◽  
Johannes Morstein ◽  
Meiyan Jin ◽  
Abrar Bhat ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Actin Cytoskeleton ◽  
Cell Fusion ◽  
Conformational Changes ◽  
Binding Motif ◽  
Mechanical Pressure ◽  
Actin Nucleation ◽  
Viral Spread ◽  
Fast Proteins ◽  
Cell Cell

Fusion-associated small transmembrane (FAST) proteins are a diverse family of nonstructural viral proteins. Once expressed on the plasma membrane of infected cells, they drive fusion with neighboring cells, increasing viral spread and pathogenicity. Unlike viral fusogens with tall ectodomains that pull two membranes together through conformational changes, FAST proteins have short fusogenic ectodomains that cannot bridge the intermembrane gap between neighboring cells. One orthoreovirus FAST protein, p14, has been shown to hijack the actin cytoskeleton to drive cell-cell fusion, but the actin adaptor-binding motif identified in p14 is not found in any other FAST protein. Here, we report that an evolutionarily divergent FAST protein, p22 from aquareovirus, also hijacks the actin cytoskeleton but does so through different adaptor proteins, Intersectin-1 and Cdc42, that trigger N-WASP–mediated branched actin assembly. We show that despite using different pathways, the cytoplasmic tail of p22 can replace that of p14 to create a potent chimeric fusogen, suggesting they are modular and play similar functional roles. When we directly couple p22 with the parallel filament nucleator formin instead of the branched actin nucleation promoting factor N-WASP, its ability to drive fusion is maintained, suggesting that localized mechanical pressure on the plasma membrane coupled to a membrane-disruptive ectodomain is sufficient to drive cell-cell fusion. This work points to a common biophysical strategy used by FAST proteins to push rather than pull membranes together to drive fusion, one that may be harnessed by other short fusogens responsible for physiological cell-cell fusion.

scholarly journals Tks5-dependent formation of circumferential podosomes/invadopodia mediates cell–cell fusion

2012 ◽  
Vol 197 (4) ◽  
pp. 553-568 ◽  
Author(s):  
Tsukasa Oikawa ◽  
Masaaki Oyama ◽  
Hiroko Kozuka-Hata ◽  
Shunsuke Uehara ◽  
Nobuyuki Udagawa ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Cell Differentiation ◽  
Cell Fusion ◽  
Underlying Mechanism ◽  
Hybrid Cells ◽  
Multinucleated Cells ◽  
Phosphoinositide 3 Kinase ◽  
Inflammatory Milieu ◽  
Invadopodia Formation ◽  
Cell Cell

Osteoclasts fuse to form multinucleated cells during osteoclastogenesis. This process is mediated by dynamic rearrangement of the plasma membrane and cytoskeleton, and it requires numerous factors, many of which have been identified. The underlying mechanism remains obscure, however. In this paper, we show that Tks5, a master regulator of invadopodia in cancer cells, is crucial for osteoclast fusion downstream of phosphoinositide 3-kinase and Src. Expression of Tks5 was induced during osteoclastogenesis, and prevention of this induction impaired both the formation of circumferential podosomes and osteoclast fusion without affecting cell differentiation. Tyrosine phosphorylation of Tks5 was attenuated in Src−/− osteoclasts, likely accounting for defects in podosome organization and multinucleation in these cells. Circumferential invadopodia formation in B16F0 melanoma cells was also accompanied by Tks5 phosphorylation. Co-culture of B16F0 cells with osteoclasts in an inflammatory milieu promoted the formation of melanoma–osteoclast hybrid cells. Our results thus reveal an unexpected link between circumferential podosome/invadopodium formation and cell–cell fusion in and beyond osteoclasts.

scholarly journals Murine Coronavirus Requires Lipid Rafts for Virus Entry and Cell-Cell Fusion but Not for Virus Release

2005 ◽  
Vol 79 (15) ◽  
pp. 9862-9871 ◽  
Author(s):  
Keum S. Choi ◽  
Hideki Aizaki ◽  
Michael M. C. Lai
Keyword(s):  
Plasma Membrane ◽  
Lipid Rafts ◽  
Cell Fusion ◽  
Lipid Raft ◽  
Virus Entry ◽  
Buoyant Density ◽  
Spike Protein ◽  
Cholesterol Depletion ◽  
Virus Binding ◽  
Cell Cell

ABSTRACT Thorp and Gallagher first reported that depletion of cholesterol inhibited virus entry and cell-cell fusion of mouse hepatitis virus (MHV), suggesting the importance of lipid rafts in MHV replication (E. B. Thorp and T. M. Gallagher, J. Virol. 78:2682-2692, 2004). However, the MHV receptor is not present in lipid rafts, and anchoring of the MHV receptor to lipid rafts did not enhance MHV infection; thus, the mechanism of lipid rafts involvement is not clear. In this study, we defined the mechanism and extent of lipid raft involvement in MHV replication. We showed that cholesterol depletion by methyl β-cyclodextrin or filipin did not affect virus binding but reduced virus entry. Furthermore, MHV spike protein bound to nonraftraft membrane at 4°C but shifted to lipid rafts at 37°C, indicating a redistribution of membrane following virus binding. Thus, the lipid raft involvement in MHV entry occurs at a step following virus binding. We also found that the viral spike protein in the plasma membrane of the infected cells was associated with lipid rafts, whereas that in the Golgi membrane, where MHV matures, was not. Moreover, the buoyant density of the virion was not changed when MHV was produced from the cholesterol-depleted cells, suggesting that MHV does not incorporate lipid rafts into the virion. These results indicate that MHV release does not involve lipid rafts. However, MHV spike protein has an inherent ability to associate with lipid rafts. Correspondingly, cell-cell fusion induced by MHV was retarded by cholesterol depletion, consistent with the association of the spike protein with lipid rafts in the plasma membrane. These findings suggest that MHV entry requires specific interactions between the spike protein and lipid rafts, probably during the virus internalization step.

scholarly journals The late endosome-resident lipid bis(monoacylglycero)phosphate is a cofactor for Lassa virus fusion

2021 ◽  
Vol 17 (9) ◽  
pp. e1009488
Author(s):  
Ruben M. Markosyan ◽  
Mariana Marin ◽  
You Zhang ◽  
Fredric S. Cohen ◽  
Gregory B. Melikyan
Keyword(s):  
Cell Fusion ◽  
Host Cells ◽  
Target Cells ◽  
Lassa Virus ◽  
Viral Glycoprotein ◽  
Glycoprotein Complex ◽  
Late Endosomes ◽  
Viral Glycoproteins ◽  
Fusion Pores ◽  
Cell Cell

Arenavirus entry into host cells occurs through a low pH-dependent fusion with late endosomes that is mediated by the viral glycoprotein complex (GPC). The mechanisms of GPC-mediated membrane fusion and of virus targeting to late endosomes are not well understood. To gain insights into arenavirus fusion, we examined cell-cell fusion induced by the Old World Lassa virus (LASV) GPC complex. LASV GPC-mediated cell fusion is more efficient and occurs at higher pH with target cells expressing human LAMP1 compared to cells lacking this cognate receptor. However, human LAMP1 is not absolutely required for cell-cell fusion or LASV entry. We found that GPC-induced fusion progresses through the same lipid intermediates as fusion mediated by other viral glycoproteins–a lipid curvature-sensitive intermediate upstream of hemifusion and a hemifusion intermediate downstream of acid-dependent steps that can be arrested in the cold. Importantly, GPC-mediated fusion and LASV pseudovirus entry are specifically augmented by an anionic lipid, bis(monoacylglycero)phosphate (BMP), which is highly enriched in late endosomes. This lipid also specifically promotes cell fusion mediated by Junin virus GPC, an unrelated New World arenavirus. We show that BMP promotes late steps of LASV fusion downstream of hemifusion–the formation and enlargement of fusion pores. The BMP-dependence of post-hemifusion stages of arenavirus fusion suggests that these viruses evolved to use this lipid as a cofactor to selectively fuse with late endosomes.

scholarly journals Plasma Membrane Integrity During Cell-Cell Fusion and in Response to Pore-Forming Drugs Is Promoted by the Penta-EF-Hand Protein PEF1 in Neurospora crassa

Genetics ◽  
2019 ◽  
pp. genetics.302363.2019 ◽  
Author(s):  
Marcel René Schumann ◽  
Ulrike Brandt ◽  
Christian Adis ◽  
Lisa Hartung ◽  
André Fleißner
Keyword(s):  
Plasma Membrane ◽  
Neurospora Crassa ◽  
Cell Fusion ◽  
Membrane Integrity ◽  
Plasma Membrane Integrity ◽  
Ef Hand ◽  
Cell Cell

scholarly journals Evolutionarily related small viral fusogens hijack distinct but modular actin nucleation pathways to drive cell-cell fusion

2020 ◽  
Author(s):  
Ka Man Carmen Chan ◽  
Ashley L. Arthur ◽  
Johannes Morstein ◽  
Meiyan Jin ◽  
Abrar Bhat ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Actin Cytoskeleton ◽  
Cell Fusion ◽  
Conformational Changes ◽  
Adaptor Proteins ◽  
Binding Motif ◽  
Mechanical Pressure ◽  
Viral Spread ◽  
Fast Proteins ◽  
Cell Cell

AbstractFusion-associated small transmembrane (FAST) proteins are a diverse family of non-structural viral proteins that, once expressed on the plasma membrane of infected cells, drive fusion with neighboring cells, increasing viral spread and pathogenicity. Unlike viral fusogens with tall ectodomains that pull two membranes together through conformational changes, FAST proteins have short fusogenic ectodomains that cannot bridge the inter-membrane gap between neighboring cells. One orthoreovirus FAST protein, p14, has been shown to hijack the actin cytoskeleton to drive cell-cell fusion, but the actin adaptor-binding motif identified in p14 is not found in any other FAST protein. Here, we report that an evolutionarily divergent FAST protein, p22 from aquareovirus, also hijacks the actin cytoskeleton but does so through different adaptor proteins, Intersectin-1 and Cdc42, that trigger N-WASP-mediated branched actin assembly. We show that despite using different pathways, the cytoplasmic tails of p22 and p14 can be exchanging to create a potent chimeric fusogen, suggesting they are modular and play similar functional roles. When we replace p22’s branched actin nucleator, N-WASP, with the parallel filament nucleator, formin, its ability to drive fusion is maintained, indicating that localized mechanical pressure on the plasma membrane coupled to a membrane-disruptive ectodomain is sufficient to drive cell-cell fusion. This work points to a common biophysical strategy used by FAST proteins to push rather than pull membranes together to drive fusion, one that may be harnessed by other short fusogens responsible for physiological cell-cell fusion.

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