Ultrastructure and behavior of actin cytoskeleton during cell wall formation in the fission yeast Schizosaccharomyces pombe

Такаги, Таичиро; Ishijima, Sanae A.; Ochi, Hisako; Osumi, Masako

doi:10.1093/jmicro/52.2.161

Cited by 47 publications

(51 citation statements)

References 37 publications

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“…Immunoelectron microscopy was performed as described previously (21,22) with anti-HA mAb (1:25 dilution) and goat antibody to mouse IgG labeled with 1-nm colloidal ultrasmall gold (1:100 dilution; GAM IgG GP-US; Aurion, Wageningen, The Netherlands). The ultra-small gold particles were enhanced by silver deposition using a silver enhancement kit (Aurion RGent SE-EM, Aurion) (20,21). The stained cells were examined using a transmission electron microscope JEM 1200 EXS (JEOL Ltd., Tokyo, Japan) at 80 kV.…”

Section: Methodsmentioning

confidence: 99%

Kre6 Protein Essential for Yeast Cell Wall β-1,6-Glucan Synthesis Accumulates at Sites of Polarized Growth

Kurita

Noda

Такаги

et al. 2011

Journal of Biological Chemistry

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Saccharomyces cerevisiae Kre6 is a type II membrane protein with amino acid sequence homology with glycoside hydrolase and is essential for ␤-1,6-glucan synthesis as revealed by the mutant phenotype, but its biochemical function is still unknown. The localization of Kre6, determined by epitope tagging, is a matter of debate. We raised anti-Kre6 rabbit antiserum and examined the localization of Kre6 and its tagged protein by immunofluorescence microscopy, subcellular fractionation in sucrose density gradients, and immunoelectron microscopy. Integration of the results indicates that the majority of Kre6 is in the endoplasmic reticulum; however, a small but significant portion is also present in the secretory vesiclelike compartments and plasma membrane. Kre6 in the latter compartments is observed as strong signals that accumulate at the sites of polarized growth by immunofluorescence. The truncated Kre6 without the N-terminal 230-amino acid cytoplasmic region did not show this polarized accumulation and had a severe defect in ␤-1,6-glucan synthesis. This is the first evidence of a ␤-1,6-glucan-related protein showing the polarized membrane localization that correlates with its biological function.

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Section: Methodsmentioning

confidence: 99%

Kre6 Protein Essential for Yeast Cell Wall β-1,6-Glucan Synthesis Accumulates at Sites of Polarized Growth

Kurita

Noda

Такаги

et al. 2011

Journal of Biological Chemistry

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“…Attempts have been made to visualize yeast actin filaments by thinsection electron microscopy (Adams and Pringle, 1984;Mulholland et al, 1994). However, actin filaments do not show up well in the crowded yeast cytoplasm, except for rare actin cables in Saccharomyces cerevisiae and pathogenic yeasts (Adams and Pringle, 1984;Kopecka et al, 2001;Yamaguchi et al, 2002), cytoplasmic "filasomes" distinct from cortical patches in fission yeast (Kanbe et al, 1989;Takagi et al, 2003), and aberrant filamentous actin structures in mutant budding yeast defective in endocytosis (Sekiya-Kawasaki et al, 2003). Immunolabeled thin sections have in some cases revealed finger-like invaginations of the plasma membrane apparently entwined in an actin helix (Mulholland et al, 1994).…”

Section: Introductionmentioning

confidence: 99%

Actin and Septin Ultrastructures at the Budding Yeast Cell Cortex

Rodal

Kozubowski

Goode

et al. 2005

MBoC

174

162

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Budding yeast has been a powerful model organism for studies of the roles of actin in endocytosis and septins in cell division and in signaling. However, the depth of mechanistic understanding that can be obtained from such studies has been severely hindered by a lack of ultrastructural information about how actin and septins are organized at the cell cortex. To address this problem, we developed rapid-freeze and deep-etch techniques to image the yeast cell cortex in spheroplasted cells at high resolution. The cortical actin cytoskeleton assembles into conical or mound-like structures composed of short, cross-linked filaments. The Arp2/3 complex localizes near the apex of these structures, suggesting that actin patch assembly may be initiated from the apex. Mutants in cortical actin patch components with defined defects in endocytosis disrupted different stages of cortical actin patch assembly. Based on these results, we propose a model for actin function during endocytosis. In addition to actin structures, we found that septin-containing filaments assemble into two kinds of higher order structures at the cell cortex: rings and ordered gauzes. These images provide the first high-resolution views of septin organization in cells. INTRODUCTIONBudding yeast has been used for over 20 yr as a model organism to study cytoskeletal function because the genes encoding many components of the cytoskeleton are conserved and because yeast cells are readily amenable to parallel genetic, biochemical, and cell biological analyses. Despite rapid advances in defining the components of the yeast cytoskeleton, its ultrastructure has been difficult to image at the resolution of the electron microscope, with the exception of microtubules (O'Toole et al., 1999). This may be due to the low abundance of the cytoskeleton in yeast relative to mammalian cells, to the presence of a cell wall that complicates extraction procedures, and to high ribosome density in the yeast cytoplasm. As a result, there has been a conspicuous deficit in our understanding of the higher order organization of actin and septin polymers in vivo (Pruyne and Bretscher, 2000). Thus, although it is possible to rapidly identify and mutate cytoskeletal proteins in yeast, there is no method by which to examine the ultrastructural consequences of these mutations. This gap could be filled by developing an approach to image the yeast cytoskeleton at high resolution.The yeast actin cytoskeleton is organized into three distinct structures: motile cortical patches that are polarized in the growing bud, actin cables that run along the motherdaughter cell axis, and a contractile cytokinetic ring at the neck of dividing cells (reviewed in Pruyne and Bretscher, 2000). Cortical actin patches contain many of the actin-associated proteins common to all eukaryotes (Goode and Rodal, 2001) and are composed of actin filaments that undergo rapid turnover (Ayscough et al., 1997). Actin is essential for yeast endocytosis (Kubler and Riezman, 1993). A number of recent studies suggest that act...

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“…Their movement can be polarized via cables with which they can associate. However, most of the patches exhibit nondirected movement, especially near the poles (Pelham and Chang, 2001;Takagi et al, 2003). Interestingly, patches seem to move away from the cell tips or the septum, rather than concentrate in these zones (Pelham and Chang, 2001).…”

Section: Actin Cytoskeleton: Structure and Dynamicsmentioning

confidence: 99%

“…Actin is also required for cell wall formation mechanisms (Osumi et al, 1998;Takagi et al, 2003). Indeed, the treatment of a protoplast with the actin polymerization inhibitor cytochalasin D or the mutation in the actin gene act1/cps8 inhibits the formation of cell wall.…”

Section: Actin Cytoskeleton: Structure and Dynamicsmentioning

confidence: 99%

“…Indeed, the treatment of a protoplast with the actin polymerization inhibitor cytochalasin D or the mutation in the actin gene act1/cps8 inhibits the formation of cell wall. Moreover, actin patches localize to initiation sites of cell wall formation in reverting protoplasts which regenerate new cell wall (Osumi et al, 1998;Takagi et al, 2003).…”

Section: Actin Cytoskeleton: Structure and Dynamicsmentioning

confidence: 99%

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Fission yeast cytoskeletons and cell polarity factors: connecting at the cortex

Carbona

Goff

2006

Biology of the Cell

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Cell polarity is a fundamental property of cells from unicellular to multicellular organisms. Most of the time, it is essential so that the cells can achieve their function. The fission yeast Schizosaccharomyces pombe is a powerful genetic model organism for studying the molecular mechanisms of the cell polarity process. Indeed, S. pombe cells are rod-shaped and cell growth is restricted at the poles. The accurate localization of the cell growth machinery at the cell cortex, which involves the actin cytoskeleton, depends on cell polarity pathways that are temporally and spatially regulated. The importance of interphase microtubules and cell polarity factors acting at the cortex of cell ends in this process has been shown. Here, we review recent advances in knowledge of molecular pathways leading to the establishment of a cellular axis in fission yeast. We also describe the role of cortical proteins and mitotic cytoskeletal rearrangements that control the symmetry of cell division.

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Ultrastructure and behavior of actin cytoskeleton during cell wall formation in the fission yeast Schizosaccharomyces pombe

Cited by 47 publications

References 37 publications

Kre6 Protein Essential for Yeast Cell Wall β-1,6-Glucan Synthesis Accumulates at Sites of Polarized Growth

Kre6 Protein Essential for Yeast Cell Wall β-1,6-Glucan Synthesis Accumulates at Sites of Polarized Growth

Actin and Septin Ultrastructures at the Budding Yeast Cell Cortex

Fission yeast cytoskeletons and cell polarity factors: connecting at the cortex

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