Chitin synthesis and localization in cell division cycle mutants of Saccharomyces cerevisiae

1983 ◽  
Vol 3 (5) ◽  
pp. 922-930
Author(s):  
R L Roberts ◽  
B Bowers ◽  
M L Slater ◽  
E Cabib
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Wall ◽  
Permissive Temperature ◽  
Wild Type ◽  
Chitin Synthesis ◽  
Specific Staining ◽  
Septal Region ◽  
Wall Components ◽  
Generalized Distribution

Growth of Saccharomyces cerevisiae cell cycle mutants cdc3, cdc4, cdc7, cdc24, and cdc28 at a nonpermissive temperature (37 degrees C) resulted in increased accumulation of chitin relative to other cell wall components, as compared with that observed at a permissive temperature (25 degrees C). Wild-type cells showed the same chitin/carbohydrate ratio at both temperatures, whereas mutants cdc13 and cdc21 yielded only a small increase in the ratio at 37 degrees C. These results confirm and extend those reported by B. F. Sloat and J. R. Pringle (Science 200:1171-1173, 1978) for mutant cdc24. The distribution of chitin in the cell wall was studied by electron microscopy, by specific staining with wheat germ agglutinin-colloidal gold complexes. At the permissive temperature, chitin was restricted to the septal region in all strains, whereas at 37 degrees C a generalized distribution of chitin in the cell wall was observed in all mutants. These results do not support a unique interdependence between the product of the cdc24 gene and localization of chitin deposition; they suggest that unbalanced conditions created in the cell by arresting the cycle at different stages result in generalized activation of the chitin synthetase zymogen. Thus, blockage of an event in the cell cycle may lead to consequences that are not functionally related to that event under normal conditions.

scholarly journals Chitin synthesis and localization in cell division cycle mutants of Saccharomyces cerevisiae.

1983 ◽  
Vol 3 (5) ◽  
pp. 922-930 ◽  
Author(s):  
R L Roberts ◽  
B Bowers ◽  
M L Slater ◽  
E Cabib
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Wall ◽  
Permissive Temperature ◽  
Wild Type ◽  
Chitin Synthesis ◽  
Specific Staining ◽  
Septal Region ◽  
Wall Components ◽  
Generalized Distribution

Growth of Saccharomyces cerevisiae cell cycle mutants cdc3, cdc4, cdc7, cdc24, and cdc28 at a nonpermissive temperature (37 degrees C) resulted in increased accumulation of chitin relative to other cell wall components, as compared with that observed at a permissive temperature (25 degrees C). Wild-type cells showed the same chitin/carbohydrate ratio at both temperatures, whereas mutants cdc13 and cdc21 yielded only a small increase in the ratio at 37 degrees C. These results confirm and extend those reported by B. F. Sloat and J. R. Pringle (Science 200:1171-1173, 1978) for mutant cdc24. The distribution of chitin in the cell wall was studied by electron microscopy, by specific staining with wheat germ agglutinin-colloidal gold complexes. At the permissive temperature, chitin was restricted to the septal region in all strains, whereas at 37 degrees C a generalized distribution of chitin in the cell wall was observed in all mutants. These results do not support a unique interdependence between the product of the cdc24 gene and localization of chitin deposition; they suggest that unbalanced conditions created in the cell by arresting the cycle at different stages result in generalized activation of the chitin synthetase zymogen. Thus, blockage of an event in the cell cycle may lead to consequences that are not functionally related to that event under normal conditions.

scholarly journals Emw1p/YNL313cp is essential for maintenance of the cell wall in Saccharomyces cerevisiae

Microbiology ◽  
2011 ◽  
Vol 157 (4) ◽  
pp. 1032-1041 ◽  
Author(s):  
Tatjana Sipling ◽  
Chao Zhai ◽  
Barry Panaretou
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Permissive Temperature ◽  
Loss Of Function ◽  
Chitin Synthesis ◽  
Calcofluor White ◽  
Compensatory Response ◽  
Protein Protein Interaction ◽  
Kinase Cascade ◽  
A Cell

There are six essential genes in the Saccharomyces cerevisiae genome which encode proteins bearing the tetratricopeptide repeat (TPR) domain that mediates protein–protein interaction. Thus far, the function of one of them, YNL313c, remains unknown. Our conditional mutants of YNL313c display osmoremedial temperature sensitivity, hypersensitivity to both Calcofluor White and low concentrations of SDS, and osmoremedial caffeine sensitivity. These are hallmarks of mutants that display cell wall defects. Accordingly we rename the gene as EMW1 (essential for maintenance of the cell wall). Loss of Emw1p function is not associated with abrogation of the cell wall integrity (CWI) MAP kinase cascade. Instead, emw1ts mutants activate this cascade even at permissive temperature, indicating that loss of Emw1p function does not cause a defect in sensors and effectors of cell wall signalling, but leads to a cell wall defect directly. Constitutive activation of the CWI cascade is reflected by the overproduction of chitin by emw1ts mutants, a compensatory response frequently displayed by cell wall mutants. Growth is restored to emw1ts mutants incubated at otherwise non-permissive temperature when GFA1 is overexpressed. GFA1 encodes the hexosephosphate aminotransferase that catalyses the rate-limiting step in the pathway that synthesizes the chitin precursor UDP-GlcNAc. The possibility that Emw1p is required for function of Gfa1p was ruled out, because the emw1ts phenotype persists when the requirement for Gfa1p is bypassed. Furthermore, if loss of Emw1p function leads to loss of function of Gfa1p, then chitin synthesis would be diminished. Instead, a stimulation of the synthesis of this polymer is detected. Consequently, the defect associated with emw1ts mutants may be associated with compromise in one of the remaining processes that depend on UDP-GlcNAc, namely N-glycosylation or glycosylphosphatidylinositol (GPI)-anchor synthesis.

scholarly journals Type 1 protein phosphatase is required for maintenance of cell wall integrity, morphogenesis and cell cycle progression in Saccharomyces cerevisiae

2000 ◽  
Vol 113 (3) ◽  
pp. 507-520 ◽  
Author(s):  
P.D. Andrews ◽  
M.J. Stark
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Wall ◽  
Actin Cytoskeleton ◽  
Map Kinase ◽  
Cell Lysis ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Bud Morphology

GLC7 encodes the catalytic subunit of type 1 protein serine/threonine phosphatase (PP1) in the yeast Saccharomyces cerevisiae. Here we have characterized the temperature-sensitive glc7-10 allele, which displays aberrant bud morphology and an abnormal actin cytoskeleton at the restrictive temperature. At 37 degrees C glc7-10 strains accumulated a high proportion of budded cells with an unmigrated nucleus, duplicated spindle pole bodies, a short spindle, delocalized cortical actin and 2C DNA content, indicating a cell cycle block prior to the metaphase to anaphase transition. glc7-10 was suppressed by growth on high osmolarity medium and exhibited temperature-sensitive cell lysis upon hypo-osmotic stress. Pkc1p, the yeast protein kinase C homolog which is thought to regulate the Mpk1p MAP kinase pathway involved in cell wall remodelling and polarized cell growth, was found to act as a dosage suppressor of glc7-10. Although neither activation of BCK1 (MEKK) by the dominant BCK1-20 mutation nor increased dosage of MKK1 (MEK) or MPK1 (MAP kinase) mimicked PKC1 as a glc7-10 dosage suppressor, extra copies of genes encoding upstream components of the Pkc1p pathway such as ROM2, RHO2, HCS77/WSC1/SLG1 and MID2 also suppressed glc7-10 effectively. Conversely, mpk1delta glc7-10 and bck1delta glc7-10 double mutants displayed a synthetic cell lysis defect compared with each single mutant and glc7-10 was hypersensitive to reduced PKC1 function, displaying highly aberrant morphologies and inviability even at the normally permissive temperature of 26 degrees C. Dephosphorylation by PP1 therefore functions positively to promote cell integrity, bud morphology and polarization of the actin cytoskeleton and glc7-10 cells require higher levels of Pkc1p activity to sustain these functions.

Yeast dom34 Mutants Are Defective in Multiple Developmental Pathways and Exhibit Decreased Levels of Polyribosomes

Genetics ◽  
1998 ◽  
Vol 149 (1) ◽  
pp. 45-56
Author(s):  
Luther Davis ◽  
JoAnne Engebrecht
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Heterologous Expression ◽  
Protein Translation ◽  
Developmental Pathways ◽  
G1 Phase ◽  
Growth Defect ◽  
Wild Type ◽  
Drosophila Homolog ◽  
Mutant Cells

Abstract The DOM34 gene of Saccharomyces cerevisiae is similar togenes found in diverse eukaryotes and archaebacteria. Analysis of dom34 strains shows that progression through the G1 phase of the cell cycle is delayed, mutant cells enter meiosis aberrantly, and their ability to form pseudohyphae is significantly diminished. RPS30A, which encodes ribosomal protein S30, was identified in a screen for high-copy suppressors of the dom34Δ growth defect. dom34Δ mutants display an altered polyribosome profile that is rescued by expression of RPS30A. Taken together, these data indicate that Dom34p functions in protein translation to promote G1 progression and differentiation. A Drosophila homolog of Dom34p, pelota, is required for the proper coordination of meiosis and spermatogenesis. Heterologous expression of pelota in dom34Δ mutants restores wild-type growth and differentiation, suggesting conservation of function between the eukaryotic members of the gene family.

scholarly journals Chs1p and Chs3p, two proteins involved in chitin synthesis, populate a compartment of the Saccharomyces cerevisiae endocytic pathway.

1996 ◽  
Vol 7 (12) ◽  
pp. 1909-1919 ◽  
Author(s):  
M Ziman ◽  
J S Chuang ◽  
R W Schekman
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Endocytic Pathway ◽  
Chitin Synthase ◽  
Cell Fractionation ◽  
Cell Wall Polysaccharide ◽  
Chitin Synthesis ◽  
A Cell ◽  
Steady State Levels ◽  
Membrane Bound

In Saccharomyces cerevisiae, the synthesis of chitin, a cell-wall polysaccharide, is temporally and spatially regulated with respect to the cell cycle and morphogenesis. Using immunological reagents, we found that steady-state levels of Chs1p and Chs3p, two chitin synthase enzymes, did not fluctuate during the cell cycle, indicating that they are not simply regulated by synthesis and degradation. Previous cell fractionation studies demonstrated that chitin synthase I activity (CSI) exists in a plasma membrane form and in intracellular membrane-bound particles called chitosomes. Chitosomes were proposed to act as a reservoir for regulated transport of chitin synthase enzymes to the division septum. We found that Chs1p and Chs3p resided partly in chitosomes and that this distribution was not cell cycle regulated. Pulse-chase cell fractionation experiments showed that chitosome production was blocked in an endocytosis mutant (end4-1), indicating that endocytosis is required for the formation or maintenance of chitosomes. Additionally, Ste2p, internalized by ligand-induced endocytosis, cofractionated with chitosomes, suggesting that these membrane proteins populate the same endosomal compartment. However, in contrast to Ste2p, Chs1p and Chs3p were not rapidly degraded, thus raising the possibility that the temporal and spatial regulation of chitin synthesis is mediated by the mobilization of an endosomal pool of chitin synthase enzymes.

scholarly journals DAF1, a mutant gene affecting size control, pheromone arrest, and cell cycle kinetics of Saccharomyces cerevisiae.

1988 ◽  
Vol 8 (11) ◽  
pp. 4675-4684 ◽  
Author(s):  
F R Cross
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Critical Size ◽  
Size Control ◽  
Mutant Gene ◽  
G1 Phase ◽  
Wild Type ◽  
Alpha Factor ◽  
Multiple Copies ◽  
Kinetics Of

The mating pheromone alpha-factor arrests Saccharomyces cerevisiae MATa cells in the G1 phase of the cell cycle. Size control is also exerted in G1, since cells do not exit G1 until they have attained a critical size. A dominant mutation (DAF1-1) which causes both alpha-factor resistance and small cell size (volume about 0.6-fold that of the wild type) has been isolated and characterized genetically and by molecular cloning. Several alpha-factor-induced mRNAs were induced equivalently in daf1+ and DAF1-1 cells. The DAF1-1 mutation consisted of a termination codon two-thirds of the way through the daf1+ coding sequence. A chromosomal deletion of DAF1 produced by gene transplacement increased cell volume about 1.5-fold; thus, DAF1-1 may be a hyperactive or deregulated allele of a nonessential gene involved in G1 size control. Multiple copies of DAF1-1 also greatly reduced the duration of the G1 phase of the cell cycle.

The lethality of p60v-src in Saccharomyces cerevisiae and the activation of p34CDC28 kinase are dependent on the integrity of the SH2 domain

1993 ◽  
Vol 105 (2) ◽  
pp. 519-528
Author(s):  
F. Boschelli ◽  
S.M. Uptain ◽  
J.J. Lightbody
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Protein Tyrosine Kinase ◽  
Kinase Activity ◽  
Cell Cycle Control ◽  
Sh2 Domain ◽  
Wild Type ◽  
Type V ◽  
Cdc28 Kinase ◽  
In Cells

The lethal effects of the expression of the oncogenic protein tyrosine kinase p60v-src in Saccharomyces cerevisiae are associated with a loss of cell cycle control at the G1/S and G2/M checkpoints. Results described here indicate that the ability of v-Src to kill yeast is dependent on the integrity of the SH2 domain, a region of the Src protein involved in recognition of proteins phosphorylated on tyrosine. Catalytically active v-Src proteins with deletions in the SH2 domain have little effect on yeast growth, unlike wild-type v-Src protein, which causes accumulation of large-budded cells, perturbation of spindle microtubules and increased DNA content when expressed. The proteins phosphorylated on tyrosine in cells expressing v-Src differ from those in cells expressing a Src protein with a deletion in the SH2 domain. Also, unlike the wild-type v-Src protein, which drastically increases histone H1-associated Cdc28 kinase activity, c-Src and an altered v-Src protein have no effect on Cdc28 kinase activity. These results indicate that the SH2 domain is functionally important in the disruption of the yeast cell cycle by v-Src.

scholarly journals Purification of profilin from Saccharomyces cerevisiae and analysis of profilin-deficient cells.

1990 ◽  
Vol 110 (1) ◽  
pp. 105-114 ◽  
Author(s):  
B K Haarer ◽  
S H Lillie ◽  
A E Adams ◽  
V Magdolen ◽  
W Bandlow ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Actin Polymerization ◽  
Yeast Cells ◽  
Predicted Amino Acid Sequence ◽  
Temperature Sensitive ◽  
Wild Type ◽  
Yeast Saccharomyces Cerevisiae ◽  
Ellipsoidal Shape ◽  
Hydrolysis Of

We have isolated profilin from yeast (Saccharomyces cerevisiae) and have microsequenced a portion of the protein to confirm its identity; the region microsequenced agrees with the predicted amino acid sequence from a profilin gene recently isolated from S. cerevisiae (Magdolen, V., U. Oechsner, G. Müller, and W. Bandlow. 1988. Mol. Cell. Biol. 8:5108-5115). Yeast profilin resembles profilins from other organisms in molecular mass and in the ability to bind to polyproline, retard the rate of actin polymerization, and inhibit hydrolysis of ATP by monomeric actin. Using strains that carry disruptions or deletions of the profilin gene, we have found that, under appropriate conditions, cells can survive without detectable profilin. Such cells grow slowly, are temperature sensitive, lose the normal ellipsoidal shape of yeast cells, often become multinucleate, and generally grow much larger than wild-type cells. In addition, these cells exhibit delocalized deposition of cell wall chitin and have dramatically altered actin distributions.

scholarly journals A Chitin Synthase with a Myosin-Like Motor Domain Is Essential for Hyphal Growth, Appressorium Differentiation, and Pathogenicity of the Maize Anthracnose Fungus Colletotrichum graminicola

2007 ◽  
Vol 20 (12) ◽  
pp. 1555-1567 ◽  
Author(s):  
Stefan Werner ◽  
Janyce A. Sugui ◽  
Gero Steinberg ◽  
Holger B. Deising
Keyword(s):  
Cell Wall ◽  
Filamentous Fungi ◽  
Hyphal Growth ◽  
Motor Domain ◽  
Plant Colonization ◽  
Colletotrichum Graminicola ◽  
Wild Type ◽  
Chitin Synthesis ◽  
Cell Wall Biogenesis ◽  
Host Cell Wall

Chitin synthesis contributes to cell wall biogenesis and is essential for invasion of solid substrata and pathogenicity of filamentous fungi. In contrast to yeasts, filamentous fungi contain up to 10 chitin synthases (CHS), which might reflect overlapping functions and indicate their complex lifestyle. Previous studies have shown that a class VI CHS of the maize anthracnose fungus Colletotrichum graminicola is essential for cell wall synthesis of conidia and vegetative hyphae. Here, we report on cloning and characterization of three additional CHS genes, CgChsI, CgChsIII, and CgChsV, encoding class I, III, and V CHS, respectively. All CHS genes are expressed during vegetative and pathogenic development. ΔCgChsI and ΔCgChsIII mutants did not differ significantly from the wild-type isolate with respect to hyphal growth and pathogenicity. In contrast, null mutants in the CgChsV gene, which encodes a CHS with an N-terminal myosin-like motor domain, are strongly impaired in vegetative growth and pathogenicity. Even in osmotically stabilized media, vegetative hyphae of ΔCgChsV mutants exhibited large balloon-like swellings, appressorial walls appeared to disintegrate during maturation, and infection cells were nonfunctional. Surprisingly, ΔCgChsV mutants were able to form dome-shaped hyphopodia that exerted force and showed host cell wall penetration rates comparable with the wild type. However, infection hyphae that formed within the plant cells exhibited severe swellings and were not able to proceed with plant colonization efficiently. Consequently, ΔCgChsV mutants did not develop macroscopically visible anthracnose disease symptoms and, thus, were nonpathogenic.

scholarly journals A Temperature-Sensitive dcw1 Mutant of Saccharomyces cerevisiae Is Cell Cycle Arrested with Small Buds Which Have Aberrant Cell Walls

2004 ◽  
Vol 3 (5) ◽  
pp. 1297-1306 ◽  
Author(s):  
Hiroshi Kitagaki ◽  
Kiyoshi Ito ◽  
Hitoshi Shimoi
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Cycle ◽  
Cell Wall ◽  
Cell Walls ◽  
Spindle Pole ◽  
Temperature Sensitive ◽  
Mrna Levels ◽  
Homologous Proteins ◽  
Aberrant Cell ◽  
Cell Wall Biogenesis

ABSTRACT Dcw1p and Dfg5p in Saccharomyces cerevisiae are homologous proteins that were previously shown to be involved in cell wall biogenesis and to be essential for growth. Dcw1p was found to be a glycosylphosphatidylinositol-anchored membrane protein. To investigate the roles of these proteins in cell wall biogenesis and cell growth, we constructed mutant alleles of DCW1 by random mutagenesis, introduced them into a Δdcw1 Δdfg5 background, and isolated a temperature-sensitive mutant, DC61 (dcw1-3 Δdfg5). When DC61 cells were incubated at 37°C, most cells had small buds, with areas less than 20% of those of the mother cells. This result indicates that DC61 cells arrest growth with small buds at 37°C. At 37°C, fewer DC61 cells had 1N DNA content and most of them still had a single nucleus located apart from the bud neck. In addition, in DC61 cells incubated at 37°C, bipolar spindles were not formed. These results indicate that DC61 cells, when incubated at 37°C, are cell cycle arrested after DNA replication and prior to the separation of spindle pole bodies. The small buds of DC61 accumulated chitin in the bud cortex, and some of them were lysed, which indicates that they had aberrant cell walls. A temperature-sensitive dfg5 mutant, DF66 (Δdcw1 dfg5-29), showed similar phenotypes. DCW1 and DFG5 mRNA levels peaked in the G1 and S phases, respectively. These results indicate that Dcw1p and Dfg5p are involved in bud formation through their involvement in biogenesis of the bud cell wall.

Sign in / Sign up

Export Citation Format

Share Document