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 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 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 Prm1p, a Pheromone-Regulated Multispanning Membrane Protein, Facilitates Plasma Membrane Fusion during Yeast Mating

2000 ◽  
Vol 151 (3) ◽  
pp. 719-730 ◽  
Author(s):  
Maxwell G. Heiman ◽  
Peter Walter
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Membrane Fusion ◽  
Cell Fusion ◽  
Plasma Membranes ◽  
Molecular Mechanisms ◽  
Transmembrane Protein ◽  
Large Fraction ◽  
Cell Contact ◽  
Yeast Mating

Cell fusion occurs throughout development, from fertilization to organogenesis. The molecular mechanisms driving plasma membrane fusion in these processes remain unknown. While yeast mating offers an excellent model system in which to study cell fusion, all genes previously shown to regulate the process act at or before cell wall breakdown; i.e., well before the two plasma membranes have come in contact. Using a new strategy in which genomic data is used to predict which genes may possess a given function, we identified PRM1, a gene that is selectively expressed during mating and that encodes a multispanning transmembrane protein. Prm1p localizes to sites of cell–cell contact where fusion occurs. In matings between Δprm1 mutants, a large fraction of cells initiate zygote formation and degrade the cell wall separating mating partners but then fail to fuse. Electron microscopic analysis reveals that the two plasma membranes in these mating pairs are tightly apposed, remaining separated only by a uniform gap of ∼8 nm. Thus, the phenotype of Δprm1 mutants defines a new step in the mating reaction in which membranes are juxtaposed, possibly through a defined adherence junction, yet remain unfused. This phenotype suggests a role for Prm1p in plasma membrane fusion.

scholarly journals Cell wall biosynthesis and the molecular mechanism of plant enlargement

2009 ◽  
Vol 36 (5) ◽  
pp. 383 ◽  
Author(s):  
John S. Boyer
Keyword(s):  
Cell Wall ◽  
Critical Level ◽  
Turgor Pressure ◽  
Chara Corallina ◽  
Wall Material ◽  
Primary Wall ◽  
Terrestrial Plants ◽  
Green Plants ◽  
Wall Deposition ◽  
Wall Growth

Recently discovered reactions allow the green alga Chara corallina (Klien ex. Willd., em. R.D.W.) to grow well without the benefit of xyloglucan or rhamnogalactan II in its cell wall. Growth rates are controlled by polygalacturonic acid (pectate) bound with calcium in the primary wall, and the reactions remove calcium from these bonds when new pectate is supplied. The removal appears to occur preferentially in bonds distorted by wall tension produced by the turgor pressure (P). The loss of calcium accelerates irreversible wall extension if P is above a critical level. The new pectate (now calcium pectate) then binds to the wall and decelerates wall extension, depositing new wall material on and within the old wall. Together, these reactions create a non-enzymatic but stoichiometric link between wall growth and wall deposition. In green plants, pectate is one of the most conserved components of the primary wall, and it is therefore proposed that the acceleration-deceleration-wall deposition reactions are of wide occurrence likely to underlie growth in virtually all green plants. C. corallina is one of the closest relatives of the progenitors of terrestrial plants, and this review focuses on the pectate reactions and how they may fit existing theories of plant growth.

Ultrastructure of a marine psychrophilic Vibrio

1970 ◽  
Vol 16 (11) ◽  
pp. 1027-1031 ◽  
Author(s):  
S. F. Kennedy ◽  
R. R. Colwell ◽  
G. B. Chapman
Keyword(s):  
Electron Microscopy ◽  
Cell Wall ◽  
Plasma Membrane ◽  
Cell Division ◽  
Wall Material ◽  
Cross Wall ◽  
Growth Stages ◽  
Culture Age ◽  
Primary Mechanism ◽  
Age Studies

The structure of Vibrio marinus strain PS-207 was studied by both phase and electron microscopy. It was found to possess a trilaminar plasma membrane and cell wall. Membrane-bounded subunits containing DNA-like material were found dispersed throughout the cytoplasm. Giant round forms or "macrospheres" were observed in all growth stages. The size, shape, and construction of the "macrospheres" showed some variation, but could not be related to culture age. Studies of cell division in V. marinus strain PS-207 indicate the primary mechanism to be a synthesis and centripetal deposition of plasma membrane with a concomitant or subsequent synthesis and centripetal deposition of cross wall material.

scholarly journals A Conserved Region in the F2 Subunit of Paramyxovirus Fusion Proteins Is Involved In Fusion Regulation

2007 ◽  
Vol 81 (15) ◽  
pp. 8303-8314 ◽  
Author(s):  
Amanda E. Gardner ◽  
Rebecca E. Dutch
Keyword(s):  
Membrane Fusion ◽  
Cell Fusion ◽  
Critical Role ◽  
Simian Virus ◽  
Hendra Virus ◽  
Heptad Repeat ◽  
F Protein ◽  
Conserved Region ◽  
Heptad Repeats ◽  
Cell Cell

ABSTRACT Paramyxoviruses utilize both an attachment protein and a fusion (F) protein to drive virus-cell and cell-cell fusion. F exists functionally as a trimer of two disulfide-linked subunits: F1 and F2. Alignment and analysis of a set of paramyxovirus F protein sequences identified three conserved blocks (CB): one in the fusion peptide/heptad repeat A domain, known to play important roles in fusion promotion, one in the region between the heptad repeats of F1 (CBF1) (A. E. Gardner, K. L. Martin, and R. E. Dutch, Biochemistry 46:5094-5105, 2007), and one in the F2 subunit (CBF2). To analyze the functions of CBF2, alanine substitutions at conserved positions were created in both the simian virus 5 (SV5) and Hendra virus F proteins. A number of the CBF2 mutations resulted in folding and expression defects. However, the CBF2 mutants that were properly expressed and trafficked had altered fusion promotion activity. The Hendra virus CBF2 Y79A and P89A mutants showed significantly decreased levels of fusion, whereas the SV5 CBF2 I49A mutant exhibited greatly increased cell-cell fusion relative to that for wild-type F. Additional substitutions at SV5 F I49 suggest that both side chain volume and hydrophobicity at this position are important in the folding of the metastable, prefusion state and the subsequent triggering of membrane fusion. The recently published prefusogenic structure of parainfluenza virus 5/SV5 F (H. S. Yin et al., Nature 439:38-44, 2006) places CBF2 in direct contact with heptad repeat A. Our data therefore indicate that this conserved region plays a critical role in stabilizing the prefusion state, likely through interactions with heptad repeat A, and in triggering membrane fusion.

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.

Mechanosensory ion channels in Chara: the influence of cell turgor pressure on touch-activated receptor potentials and action potentials

2001 ◽  
Vol 28 (7) ◽  
pp. 551 ◽  
Author(s):  
Virginia A. Shepherd ◽  
Teruo Shimmen ◽  
Mary J. Beilby
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Action Potentials ◽  
Turgor Pressure ◽  
Membrane Cytoskeleton ◽  
Stimulus Energy ◽  
Cell Turgor ◽  
Touch Response ◽  
Solution Changes

Chara cells produce receptor potentials (RPDs) in response to mechanical stimulation. We have used a mechanostimulatory device to compare characteristics of touch-activated RPDs and action potentials (APs) when cell turgor pressure was changed. The device delivered a series of mechanical stimulations of increasing energy (F0.5, F1, F2, F3, F4, F5 and F6). Cells were alternately stimulated in artificial pondwater (APW) and a sorbitol series, in long-term experiments, involving up to six solution changes. The calculated cell turgor pressures were about 0.6 MPa (APW), and 0.49 MPa, 0.37 MPa, 0.24 MPa and 0.12 MPa in 50, 100, 150 and 200 mM sorbitol–APW, respectively. In other experiments, cells were pre-conditioned in the sorbitol solutions, and then transferred to APW. All cells were allowed long recovery periods (40–60 min) after APs or solution transfers. Only small changes in cell conductance were observed in I–V and G–V analysis of unstimulated cells after reducing turgor pressure from 0.59 MPa to 0.24 MPa. In APW, the RPDs increased in amplitude and duration with increased stimulus energy until the threshold RPD was reached, and an AP was triggered, usually between stimulus F4 and F5. Cells with decreased turgor pressure became more sensitive to stimulation, giving threshold RPDs or APs with smaller stimulus (e.g. between F0.5 and F3). Conversely, an increase in cell turgor pressure (return to APW) led to a decrease in sensitivity to stimulus. When turgor pressure was greatly decreased (to 0.12 MPa), some cells became unresponsive or gave unusual responses. However, only the mechanical part of the touch response was affected by changing the cell turgor pressure. The mean amplitudes of the subthreshold and threshold RPD (that triggers the AP), and of the touch-activated APs, were independent of cell turgor pressure, although action potentials had smaller amplitude when turgor was reduced to about 0.12 MPa. The amplitude of the subthreshold RPD was close to 20 mV, and the amplitude of the threshold RPD was close to 50 mV, in all cells. If tension of the cell wall–plasma membrane–cytoskeleton complex decreased along with decreased cell turgor pressure, a given stimulus could stretch the complex to a greater extent, resulting in activation of more mechanosensory channels. The effect on the RPD of changes in cell turgor pressure is discussed in relation to the mechanical properties of the cell wall–plasma membrane–cytoskeleton complex.

scholarly journals Osmotic Balance Regulates Cell Fusion during Mating in Saccharomyces cerevisiae

1997 ◽  
Vol 138 (5) ◽  
pp. 961-974 ◽  
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.

Cell wall structures of Epicoccum nigrum (Hyphomycetes)

1973 ◽  
Vol 51 (5) ◽  
pp. 1071-1073 ◽  
Author(s):  
J. A. Brushaber ◽  
R. H. Haskins
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Outer Layer ◽  
Secondary Wall ◽  
Outer Wall ◽  
Wall Layer ◽  
Wall Material ◽  
Primary Wall ◽  
Electron Dense Material ◽  
Wall Structures

Two structurally distinct types of secondary wall layers are present in older hyphae in addition to the primary wall. A coarsely fibrous outer wall layer often becomes quite massive and frequently fuses with the outer wall layers of adjacent cells in the formation of hyphal strands. The uneven deposition of this outer layer often produces large verrucosities. The inner secondary wall layer is relatively electron transparent and contains a reticulum of electron-dense lines. The interface of the inner secondary wall with the cytoplasm is often very irregular, and sections of the plasma membrane are frequently overlain by wall material. The outer secondary wall of conidia is composed of an electron-dense material different from that of the outer wall of hyphae. Cells in the multicellular conidia tend to be polyhedral in shape with either very thick primary walls or thin primary walls having a thick inner wall deposit.

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