Dataset Information

Differential gene expression signatures for cell wall integrity found in chitin synthase II (chs2Delta) and myosin II (myo1Delta) deficient cytokinesis mutants of Saccharomyces cerevisiae.

ABSTRACT:

Background

Myosin II-dependent contraction of the cytokinetic ring and primary septum formation by chitin synthase II are interdependent processes during cytokinesis in Saccharomyces cerevisiae. Hence, null mutants of myosin II (myo1Delta) and chitin synthase II (chs2Delta) share multiple morphological and molecular phenotypes. To understand the nature of their interdependent functions, we will seek to identify genes undergoing transcriptional regulation in chs2Delta strains and to establish a transcription signature profile for comparison with myo1Delta strains.

Results

A total of 467 genes were commonly regulated between myo1Delta and chs2Delta mutant strains (p ConclusionComparison of the chs2Delta and myo1Delta gene expression profiles revealed similarities in the biological process categories that respond to the chs2Delta and myo1Delta gene mutations. This supports the view that these mutations affect a common function in cytokinesis. Despite their similarities, these mutants exhibited significant differences in expression of the main transcriptional fingerprint for cell wall stress and their requirement of the CWIP for survival.

SUBMITTER: Rodriguez-Quinones JF

PROVIDER: S-EPMC2684100 | biostudies-literature | 2009 May

REPOSITORIES: biostudies-literature

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Publications

Differential gene expression signatures for cell wall integrity found in chitin synthase II (chs2Delta) and myosin II (myo1Delta) deficient cytokinesis mutants of Saccharomyces cerevisiae.

Rodríguez-Quiñones José F JF Rodríguez-Medina José R JR

BMC research notes 20090509

<h4>Background</h4>Myosin II-dependent contraction of the cytokinetic ring and primary septum formation by chitin synthase II are interdependent processes during cytokinesis in Saccharomyces cerevisiae. Hence, null mutants of myosin II (myo1Delta) and chitin synthase II (chs2Delta) share multiple morphological and molecular phenotypes. To understand the nature of their interdependent functions, we will seek to identify genes undergoing transcriptional regulation in chs2Delta strains and to estab ...[more]

PMID: 19426543

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Project description:BackgroundThe Saccharomyces cerevisiae MYO1 gene encodes the myosin II heavy chain (Myo1p), a protein required for normal cytokinesis in budding yeast. Myo1p deficiency in yeast (myo1Delta) causes a cell separation defect characterized by the formation of attached cells, yet it also causes abnormal budding patterns, formation of enlarged and elongated cells, increased osmotic sensitivity, delocalized chitin deposition, increased chitin synthesis, and hypersensitivity to the chitin synthase III inhibitor Nikkomycin Z. To determine how differential expression of genes is related to these diverse cell wall phenotypes, we analyzed the global mRNA expression profile of myo1Delta strains.ResultsGlobal mRNA expression profiles of myo1Delta strains and their corresponding wild type controls were obtained by hybridization to yeast oligonucleotide microarrays. Results for selected genes were confirmed by real time RT-PCR. A total of 547 differentially expressed genes (p < or = 0.01) were identified with 263 up regulated and 284 down regulated genes in the myo1Delta strains. Gene set enrichment analysis revealed the significant over-representation of genes in the protein biosynthesis and stress response categories. The SLT2/MPK1 gene was up regulated in the microarray, and a myo1Deltaslt2Delta double mutant was non-viable. Overexpression of ribosomal protein genes RPL30 and RPS31 suppressed the hypersensitivity to Nikkomycin Z and increased the levels of phosphorylated Slt2p in myo1Delta strains. Increased levels of phosphorylated Slt2p were also observed in wild type strains under these conditions.ConclusionFollowing this analysis of global mRNA expression in yeast myo1Delta strains, we conclude that 547 genes were differentially regulated in myo1Delta strains and that the stress response and protein biosynthesis gene categories were coordinately regulated in this mutant. The SLT2/MPK1 gene was confirmed to be essential for myo1Delta strain viability, supporting that the up regulated stress response genes are regulated by the PKC1 cell integrity pathway. Suppression of Nikkomycin Z hypersensitivity together with Slt2p phosphorylation was caused by the overexpression of ribosomal protein genes RPL30 and RPS31. These ribosomal protein mRNAs were down regulated in the myo1Delta arrays, suggesting that down regulation of ribosomal biogenesis may affect cell integrity in myo1Delta strains.

Project description:Just before bud emergence, a Saccharomyces cerevisiae cell forms a ring of chitin in its cell wall; this ring remains at the base of the bud as the bud grows and ultimately forms part of the bud scar marking the division site on the mother cell. The chitin ring seems to be formed largely or entirely by chitin synthase III, one of the three known chitin synthases in S. cerevisiae. The chitin ring does not form normally in temperature-sensitive mutants defective in any of four septins, a family of proteins that are constituents of the "neck filaments" that lie immediately subjacent to the plasma membrane in the mother-bud neck. In addition, a synthetic-lethal interaction was found between cdc12-5, a temperature-sensitive septin mutation, and a mutant allele of CHS4, which encodes an activator of chitin synthase III. Two-hybrid analysis revealed no direct interaction between the septins and Chs4p but identified a novel gene, BNI4, whose product interacts both with Chs4p and Cdc10p and with one of the septins, Cdc10p; this analysis also revealed an interaction between Chs4p and Chs3p, the catalytic subunit of chitin synthase III. Bni4p has no known homologues; it contains a predicted coiled-coil domain, but no other recognizable motifs. Deletion of BNI4 is not lethal, but causes delocalization of chitin deposition and aberrant cellular morphology. Overexpression of Bni4p also causes delocalization of chitin deposition and produces a cellular morphology similar to that of septin mutants. Immunolocalization experiments show that Bni4p localizes to a ring at the mother-bud neck that lies predominantly on the mother-cell side (corresponding to the predominant site of chitin deposition). This localization depends on the septins but not on Chs4p or Chs3p. A GFP-Chs4p fusion protein also localizes to a ring at the mother-bud neck on the mother-cell side. This localization is dependent on the septins, Bni4p, and Chs3p. Chs3p, whose normal localization is similar to that of Chs4p, does not localize properly in bni4, chs4, or septin mutant strains or in strains that accumulate excess Bni4p. In contrast, localization of the septins is essentially normal in bni4, chs4, and chs3 mutant strains and in strains that accumulate excess Bni4p. Taken together, these results suggest that the normal localization of chitin synthase III activity is achieved by assembly of a complex in which Chs3p is linked to the septins via Chs4p and Bni4p.

Project description:Penicillium digitatum is the main postharvest pathogen of citrus fruit and is responsible for important economic losses in spite of the massive use of fungicides. The fungal cell wall (CW) and its specific component chitin are potential targets for the development of new antifungal molecules. Among these are the antifungal peptides and proteins that specifically interact with fungal CW. Chitin is synthesized by a complex family of chitin synthases (Chs), classified into up to eight classes within three divisions. Previously, we obtained and characterized a mutant of P. digitatum in the class VII gene (ΔchsVII), which contains a short myosin motor-like domain (MMD). In this report, we extend our previous studies to the characterization of mutants in chsII and in the gene coding for the other MMD-Chs (chsV), and study the role of chitin synthases in the sensitivity of P. digitatum to the self-antifungal protein AfpB, and to AfpA obtained from P. expansum. The ΔchsII mutant showed no significant phenotypic and virulence differences with the wild type strain, except in the production and morphology of the conidia. In contrast, mutants in chsV showed a more dramatic phenotype than the previous ΔchsVII, with reduced growth and conidial production, increased chitin content, changes in mycelial morphology and a decrease in virulence to citrus fruit. Mutants in chsVII were specifically more tolerant than the wild type to nikkomycin Z, an antifungal inhibitor of chitin biosynthesis. Treatment of P. digitatum with its own antifungal protein AfpB resulted in an overall reduction in the expression of the chitin synthase genes. The mutants corresponding to MMD chitin synthases exhibited differential sensitivity to the antifungal proteins AfpA and AfpB, ΔchsVII being more susceptible than its parental strain and ΔchsV being slightly more tolerant despite its reduced growth in liquid broth. Taking these results together, we conclude that the MMD-containing chitin synthases affect cell wall integrity and sensitivity to antifungal proteins in P. digitatum.

Dataset Information

Differential gene expression signatures for cell wall integrity found in chitin synthase II (chs2Delta) and myosin II (myo1Delta) deficient cytokinesis mutants of Saccharomyces cerevisiae.

Background

Results

Publications

Differential gene expression signatures for cell wall integrity found in chitin synthase II (chs2Delta) and myosin II (myo1Delta) deficient cytokinesis mutants of Saccharomyces cerevisiae.

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