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Characterization of cwl1+, a gene from Schizosaccharomyces pombe whose overexpression causes cell lysis
1Instituto de Microbiologia Bioquimica, Consejo Superior de Investigaciones Cientificas, Salamanca, Spain.
Abstract:
From a Schizosaccharomyces pombe genomic library we have isolated the gene cwl1+ that causes cell lysis when it is overexpressed in the absence of an osmotic stabilizer. Southern hybridization showed that cwl1+ exists as a single copy in the S. pombe genome. The cwl1+ gene nucleotide sequence revealed a putative open reading frame of 924 bp encoding a polypeptide of 308 amino acids with a calculated Mt of 27000. The cwl1+ DNA hybridizes to a major RNA transcript of 1.5 kb whose 5' end maps at a position 452 bp upstream from the predicted translation start. Comparison of the amino acid sequence with those included in the current databases, showed no significant similarity to any known sequences. Cells overexpressing the cwl1+ gene under the control of the S. pombe nmt inducible promoter displayed a reduced cell wall content, were unable to separate after division and lysed drastically in the absence of osmotic stabilizer. Disruption of the cwl1+ gene caused no noticeable phenotype.
Insights
We identified the Schizosaccharomyces pombe gene cwl1+, which causes cell lysis when overexpressed without osmotic support. Disrupting this gene had no observable effect, suggesting a non-essential role in normal cell growth.
Area of Science:
- * Molecular Biology
- * Genetics
- * Cell Biology
Background:
- * Understanding gene function in model organisms like Schizosaccharomyces pombe is crucial for deciphering fundamental biological processes.
- * Cell wall integrity is essential for yeast survival, and genes regulating this process are key targets for research.
Purpose of the Study:
- * To isolate and characterize a novel gene from Schizosaccharomyces pombe involved in cell wall maintenance.
- * To investigate the function of the isolated gene, designated cwl1+, through overexpression and gene disruption studies.
Main Methods:
- * Screening of a Schizosaccharomyces pombe genomic library for genes causing cell lysis upon overexpression.
- * Nucleotide sequencing of the cwl1+ gene and analysis of its predicted protein product.
- * Overexpression studies using the nmt inducible promoter and gene disruption experiments.
Main Results:
- * Isolation of the cwl1+ gene, which, when overexpressed, leads to cell lysis in the absence of osmotic stabilizers.
- * Sequence analysis revealed a 308-amino acid polypeptide with no significant similarity to known proteins.
- * Overexpression resulted in reduced cell wall content, impaired cell separation, and drastic lysis, while cwl1+ gene disruption showed no phenotype.
Conclusions:
- * The cwl1+ gene product plays a role in maintaining cell wall integrity in Schizosaccharomyces pombe, particularly under stress conditions.
- * Overexpression of cwl1+ disrupts cell wall synthesis and cell division, leading to cell death.
- * The lack of a phenotype upon gene disruption suggests functional redundancy or a role only under specific stress conditions.