In budding yeast, contraction of the actomyosin ring and formation of the primary septum at cytokinesis depend on each other

2002 ◽  
Vol 115 (2) ◽  
pp. 293-302 ◽  
Author(s):  
Martin Schmidt ◽  
Blair Bowers ◽  
Archana Varma ◽  
Dong-Hyun Roh ◽  
Enrico Cabib
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Double Mutant ◽  
Chitin Synthase ◽  
Contractile Ring ◽  
Ring Contraction ◽  
Primary Septum ◽  
Common Process ◽  
Lateral Walls

Saccharomyces cerevisiae chs2 mutants are unable to synthesize primary septum chitin, and myo1 mutants cannot construct a functional contractile ring. The morphology of the two mutants, as observed by electron microscopy, is very similar. In both cases, neither an invagination of the plasma membrane, which normally results from contraction of the actomyosin ring, nor generation of a chitin disc, the primary septum, is observed. Rather, both mutants are able to complete cytokinesis by an abnormal process in which lateral walls thicken gradually and finally meet over an extended region, giving rise to a thick septum lacking the normal trilaminar structure and often enclosing lacunae. Defects in chs2 or myo1 strains were not aggravated in a double mutant, an indication that the corresponding proteins participate in a common process. In contrast, in a chs3 background the chs2 mutation is lethal and the myo1 defect is greatly worsened, suggesting that the synthesis of chitin catalyzed by chitin synthase III is necessary for the functionality of the remedial septa. Both chs2 and myo1 mutants show abnormalities in budding pattern and a decrease in the level of certain proteins associated with budding, such as Bud3p, Bud4p and Spa2p. The possible reasons for these phenotypes and for the interdependence between actomyosin ring contraction and primary septum formation are discussed.

scholarly journals Role of Inn1 and its interactions with Hof1 and Cyk3 in promoting cleavage furrow and septum formation in S. cerevisiae

2009 ◽  
Vol 185 (6) ◽  
pp. 995-1012 ◽  
Author(s):  
Ryuichi Nishihama ◽  
Jennifer H. Schreiter ◽  
Masayuki Onishi ◽  
Elizabeth A. Vallen ◽  
Julia Hanna ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mitotic Exit ◽  
Chitin Synthase ◽  
Cleavage Furrow ◽  
Sh3 Domains ◽  
C2 Domains ◽  
Division Site ◽  
Primary Septum ◽  
N Terminus

Cytokinesis requires coordination of actomyosin ring (AMR) contraction with rearrangements of the plasma membrane and extracellular matrix. In Saccharomyces cerevisiae, new membrane, the chitin synthase Chs2 (which forms the primary septum [PS]), and the protein Inn1 are all delivered to the division site upon mitotic exit even when the AMR is absent. Inn1 is essential for PS formation but not for Chs2 localization. The Inn1 C-terminal region is necessary for localization, and distinct PXXP motifs in this region mediate functionally important interactions with SH3 domains in the cytokinesis proteins Hof1 (an F-BAR protein) and Cyk3 (whose overexpression can restore PS formation in inn1Δ cells). The Inn1 N terminus resembles C2 domains but does not appear to bind phospholipids; nonetheless, when overexpressed or fused to Hof1, it can provide Inn1 function even in the absence of the AMR. Thus, Inn1 and Cyk3 appear to cooperate in activating Chs2 for PS formation, which allows coordination of AMR contraction with ingression of the cleavage furrow.

scholarly journals Altered Na+ and Li+Homeostasis in Saccharomyces cerevisiae Cells Expressing the Bacterial Cation Antiporter NhaA

1998 ◽  
Vol 180 (12) ◽  
pp. 3131-3136 ◽  
Author(s):  
Roc Ros ◽  
Consuelo Montesinos ◽  
Abraham Rimon ◽  
Etana Padan ◽  
Ramón Serrano
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Double Mutant ◽  
Vacuolar Membrane ◽  
Intracellular Sodium ◽  
Intracellular Pool ◽  
Sodium Accumulation ◽  
Sodium Tolerance ◽  
Yeast Mutants ◽  
In Cells

ABSTRACT The bacterial Na+(Li+)/H+antiporter NhaA has been expressed in the yeast Saccharomyces cerevisiae. NhaA was present in both the plasma membrane and internal membranes, and it conferred lithium but not sodium tolerance. In cells containing the yeast Ena1-4 (Na+, Li+) extrusion ATPase, the extra lithium tolerance conferred by NhaA was dependent on a functional vacuolar H+ ATPase and correlated with an increase of lithium in an intracellular pool which exhibited slow efflux of cations. In yeast mutants without (Na+, Li+) ATPase, lithium tolerance conferred by NhaA was not dependent on a functional vacuolar H+ ATPase and correlated with a decrease of intracellular lithium. NhaA was able to confer sodium tolerance and to decrease intracellular sodium accumulation in a double mutant devoid of both plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase. These results indicate that the bacterial antiporter NhaA expressed in yeast is functional at both the plasma membrane and the vacuolar membrane. The phenotypes conferred by its expression depend on the functionality of plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase.

scholarly journals A Method of Isolating Anucleated Yeast Protoplasts Unable to Synthesize the Glucan Fibrillar Component of the Wall

Microbiology ◽  
2000 ◽  
Vol 81 (1) ◽  
pp. 111-120 ◽  
Author(s):  
Marie Kopecká ◽  
M. Gabriel ◽  
O. Nečas
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Protein Synthesis ◽  
Plasma Membrane ◽  
Light And Electron Microscopy ◽  
Sharp Contrast ◽  
Yeast Protoplasts ◽  
Inhibition Of Protein Synthesis ◽  
Reproducible Method ◽  
Fibrillar Component

A mixture of nucleated and anucleated protoplasts was produced from log-phase Saccharomyces cerevisiae by the use of snail enzymes. The mixture was separated by centrifugation, and anucleated protoplasts were studied by means of light and electron microscopy. Anucleated protoplasts did not synthesize glucan fibrils even though they seemed to contain all other basic structures in their cytoplasm, and the structure of the plasma membrane was unchanged. This was in sharp contrast to ordinary nucleated protoplasts which synthesized glucan fibrils even after inhibition of protein synthesis by cycloheximide. The reason for this behaviour of anucleated protoplasts is not clear. Such anucleated yeast protoplasts represent the first example of uniform anucleated fungi produced by a reproducible method.

scholarly journals Chs6p-dependent Anterograde Transport of Chs3p from the Chitosome to the Plasma Membrane inSaccharomyces cerevisiae

1998 ◽  
Vol 9 (6) ◽  
pp. 1565-1576 ◽  
Author(s):  
Michael Ziman ◽  
John S. Chuang ◽  
Michael Tsung ◽  
Susan Hamamoto ◽  
Randy Schekman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Cell Surface ◽  
Cell Separation ◽  
Secretory Pathway ◽  
Protein Transport ◽  
Subcellular Fractionation ◽  
Chitin Synthase ◽  
Anterograde Transport ◽  
Membrane Compartment

Chitin synthase III (CSIII), an enzyme required to form a chitin ring in the nascent division septum of Saccharomyces cerevisiae, may be transported to the cell surface in a regulated manner. Chs3p, the catalytic subunit of CSIII, requires the product of CHS6 to be transported to or activated at the cell surface. We find that chs6Δ strains have morphological abnormalities similar to those of chs3mutants. Subcellular fractionation and indirect immunofluorescence indicate that Chs3p distribution is altered in chs6mutant cells. Order-of-function experiments usingend4–1 (endocytosis-defective) and chs6mutants indicate that Chs6p is required for anterograde transport of Chs3p from an internal endosome-like membrane compartment, the chitosome, to the plasma membrane. As a result, chs6strains accumulate Chs3p in chitosomes. Chs1p, a distinct chitin synthase that acts during or after cell separation, is transported normally in chs6 mutants, suggesting that Chs1p and Chs3p are independently packaged during protein transport through the late secretory pathway.

scholarly journals The localization of chitin synthase mediates the patterned deposition of chitin in developing Drosophila bristles

2019 ◽  
Author(s):  
Paul N. Adler
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Complex Structure ◽  
Chitin Synthase ◽  
Cell Type ◽  
Body Regions ◽  
Transmission Electron ◽  
Successful Group ◽  
Sensory Bristles ◽  
Parallel Arrays

AbstractThe insect exoskeleton is a complex structure that is a key for the life style of this very successful group of animals. It contains proteins, lipids and the N-acetyl glucosamine polymer chitin. Chitin is synthesized by the enzyme chitin synthase. In most body regions, chitin fibrils are found in a stack of parallel arrays that can be detected by transmission electron microscopy. Each array is rotated with respect to the layers above and below. In sensory bristles, chitin primarily accumulates in bands parallel to the proximal/distal axis of the bristle. These bands are visible by confocal microscopy providing experimental advantages. We have used this cell type and an edited chitin synthase gene to establish that the bands of chitin are closely associated with stripes of chitin synthase, arguing the localization of chitin synthase plays an important role in mediating the patterned deposition of chitin. This is reminiscent of what has been seen for chitin and chitin synthase in fungi and between cellulose and cellulose synthase in plants. Several genes are known to be essential for proper chitin deposition. We found one of these, Rab11 is required for the insertion of chitin synthase into the plasma membrane and a second, duskylike is required for plasma membrane chitin synthase to localize properly into stripes. We also established that the actin cytoskeleton is required for the proper localization of chitin synthase and chitin in developing sensory bristles.

scholarly journals The plasma membrane ferrireductase activity of Saccharomyces cerevisiae is partially controlled by cyclic AMP

1991 ◽  
Vol 280 (2) ◽  
pp. 545-548 ◽  
Author(s):  
E Lesuisse ◽  
B Horion ◽  
P Labbe ◽  
F Hilger
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Cyclic Amp ◽  
Double Mutant ◽  
Synthetic Medium ◽  
Iron Limitation ◽  
Permissive Temperature ◽  
Extract Content ◽  
Dependent Protein ◽  
Membrane Bound

The plasma-membrane-bound ferrireductase activity of ras1 and ras2 mutants of Saccharomyces cerevisiae is not induced in response to iron limitation. This phenotype was suppressed by the bcy1 mutation in ras2 but not in ras1 mutants. The cellular haem content of ras-1-bearing strains decreased dramatically when cells were grown in semi-synthetic medium (low yeast extract content), which could account for their very low ferrireductase activity. The ferrireductase activity of cdc25 and cdc35 mutants dropped when the cells were shifted to a non-permissive temperature. This drop was prevented in the double mutant cdc35 sra5 by adding cyclic AMP to the growth medium. We propose that ferrireductase activity is under the control of a cyclic AMP-dependent protein phosphorylation.

scholarly journals Vesicles carry most exocyst subunits to exocytic sites marked by the remaining two subunits, Sec3p and Exo70p

2004 ◽  
Vol 167 (5) ◽  
pp. 889-901 ◽  
Author(s):  
Charles Boyd ◽  
Thom Hughes ◽  
Marc Pypaert ◽  
Peter Novick
Keyword(s):  
Electron Microscopy ◽  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Immunogold Electron Microscopy ◽  
Secretory Vesicles ◽  
Actin Assembly ◽  
Yeast Saccharomyces Cerevisiae ◽  
Family Protein ◽  
Photobleaching Recovery ◽  
Discrete Domains

Exocytosis in the budding yeast Saccharomyces cerevisiae occurs at discrete domains of the plasma membrane. The protein complex that tethers incoming vesicles to sites of secretion is known as the exocyst. We have used photobleaching recovery experiments to characterize the dynamic behavior of the eight subunits that make up the exocyst. One subset (Sec5p, Sec6p, Sec8p, Sec10p, Sec15p, and Exo84p) exhibits mobility similar to that of the vesicle-bound Rab family protein Sec4p, whereas Sec3p and Exo70p exhibit substantially more stability. Disruption of actin assembly abolishes the ability of the first subset of subunits to recover after photobleaching, whereas Sec3p and Exo70p are resistant. Immunogold electron microscopy and epifluorescence video microscopy indicate that all exocyst subunits, except for Sec3p, are associated with secretory vesicles as they arrive at exocytic sites. Assembly of the exocyst occurs when the first subset of subunits, delivered on vesicles, joins Sec3p and Exo70p on the plasma membrane. Exocyst assembly serves to both target and tether vesicles to sites of exocytosis.

scholarly journals Extracellular cell wall β(1,3)glucan is required to couple septation to actomyosin ring contraction

2013 ◽  
Vol 203 (2) ◽  
pp. 265-282 ◽  
Author(s):  
Javier Muñoz ◽  
Juan Carlos G. Cortés ◽  
Matthias Sipiczki ◽  
Mariona Ramos ◽  
José Angel Clemente-Ramos ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Cell Wall ◽  
Plasma Membrane ◽  
Animal Cells ◽  
Contractile Ring ◽  
Septum Formation ◽  
Main Cell ◽  
Primary Septum ◽  
Pulling Forces

Cytokinesis has been extensively studied in different models, but the role of the extracellular cell wall is less understood. Here we studied this process in fission yeast. The essential protein Bgs4 synthesizes the main cell wall β(1,3)glucan. We show that Bgs4-derived β(1,3)glucan is required for correct and stable actomyosin ring positioning in the cell middle, before the start of septum formation and anchorage to the cell wall. Consequently, β(1,3)glucan loss generated ring sliding, oblique positioned rings and septa, misdirected septum synthesis indicative of relaxed rings, and uncoupling between a fast ring and membrane ingression and slow septum synthesis, suggesting that cytokinesis can progress with defective septum pushing and/or ring pulling forces. Moreover, Bgs4-derived β(1,3)glucan is essential for secondary septum formation and correct primary septum completion. Therefore, our results show that extracellular β(1,3)glucan is required for cytokinesis to connect the cell wall with the plasma membrane and for contractile ring function, as proposed for the equivalent extracellular matrix in animal cells.

Electron microscopy of endocardial cell populations

1978 ◽  
Vol 36 (2) ◽  
pp. 486-487
Author(s):  
T. G. Sarphie ◽  
C. R. Comer ◽  
D. J. Allen
Keyword(s):  
Electron Microscopy ◽  
Plasma Membrane ◽  
Cellular Transformation ◽  
Cell Populations ◽  
Cell Surfaces ◽  
Kb Cells ◽  
Dna Viruses ◽  
Cell Cycles ◽  
Ultrastructural Studies ◽  
Endocardial Cell

Previous ultrastructural studies have characterized surface morphology during norma cell cycles in an attempt to associate specific changes with specific metabolic processes occurring within the cell. It is now known that during the synthetic ("S") stage of the cycle, when DNA and other nuclear components are synthesized, a cel undergoes a doubling in volume that is accompanied by an increase in surface area whereby its plasma membrane is elaborated into a variety of processes originally referred to as microvilli. In addition, changes in the normal distribution of glycoproteins and polysaccharides derived from cell surfaces have been reported as depreciating after cellular transformation by RNA or DNA viruses and have been associated with the state of growth, irregardless of the rate of proliferation. More specifically, examination of the surface carbohydrate content of synchronous KB cells were shown to be markedly reduced as the cell population approached division Comparison of hamster kidney fibroblasts inhibited by vinblastin sulfate while in metaphase with those not in metaphase demonstrated an appreciable decrease in surface carbohydrate in the former.

Triple Fixation For Electron Microscopy

1968 ◽  
Vol 26 ◽  
pp. 90-91 ◽  
Author(s):  
M. A. Hayat
Keyword(s):  
Electron Microscopy ◽  
Endoplasmic Reticulum ◽  
Electron Beam ◽  
Plasma Membrane ◽  
Myelin Sheath ◽  
Good Deal ◽  
Granular Structure ◽  
Oxidizing Agent ◽  
Fixation Technique ◽  
Large Clusters

Potassium permanganate has been successfully employed to study membranous structures such as endoplasmic reticulum, Golgi, plastids, plasma membrane and myelin sheath. Since KMnO4 is a strong oxidizing agent, deposition of manganese or its oxides account for some of the observed contrast in the lipoprotein membranes, but a good deal of it is due to the removal of background proteins either by dehydration agents or by volatalization under the electron beam. Tissues fixed with KMnO4 exhibit somewhat granular structure because of the deposition of large clusters of stain molecules. The gross arrangement of membranes can also be modified. Since the aim of a good fixation technique is to preserve satisfactorily the cell as a whole and not the best preservation of only a small part of it, a combination of a mixture of glutaraldehyde and acrolein to obtain general preservation and KMnO4 to enhance contrast was employed to fix plant embryos, green algae and fungi.

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