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Saccharomyces cerevisiae gene ISW2 encodes a microtubule-interacting protein required for premeiotic DNA replication
P Trachtulcová1, I Janatová, S D Kohlwein
1Institute of Microbiology, Academy of Sciences of the Czech Republic, 14220 Prague, Czech Republic.
Abstract:
A molecular genetic characterization of the ORF YOR304W (ISW2), identified in a screen of a yeast lambdagt11 library using a monoclonal antibody that reacts with a 210 kDa mammalian microtubule-interacting protein, is presented in this paper. The protein encoded by the ORF YOR304W is 50% identical to the Drosophila nucleosome remodelling factor ISWI and is therefore a new member of the SNF2 protein family and has been recently entered into SDG as ISW2. Although not essential for vegetative growth, we found that the ISW2 gene is required for early stages in sporulation. The isw2 homozygous deletant diploid strain was blocked in the G(1) phase of the cell cycle, unable to execute the premeiotic DNA replication and progress through the nuclear meiotic division cycle. ISW2 expression from a multicopy plasmid had the same effect as deletion, but ISW2 expression from a centromeric plasmid rescued the deletion phenotype. In vegetatively growing diploid cells, the Isw2 protein was preferentially found in the cytoplasm, co-localizing with microtubules. An accumulation of the Isw2 protein within the nucleus was observed in cells entering sporulation. Together with data published very recently by Tsukiyama et al. (1999), we propose a role for the Isw2 protein in facilitating chromatin accessibility for transcriptional factor(s) that positively regulate meiosis/sporulation-specific genes.
Insights
The ISW2 gene is crucial for yeast sporulation, regulating cell cycle progression and DNA replication. Its protein product, Isw2, facilitates chromatin accessibility for genes involved in meiosis.
Area of Science:
- Molecular Biology
- Yeast Genetics
- Cell Cycle Regulation
Background:
- The ISW2 gene, encoding a protein homologous to Drosophila ISWI, was identified via antibody screening.
- ISW2 belongs to the SNF2 family of proteins involved in chromatin remodeling.
Purpose of the Study:
- To molecularly and genetically characterize the yeast ISW2 gene and its encoded protein.
- To investigate the role of ISW2 in yeast cell cycle progression, specifically during sporulation.
Main Methods:
- Molecular cloning and genetic manipulation of the ISW2 gene in yeast (Saccharomyces cerevisiae).
- Analysis of cell cycle progression using flow cytometry.
- Subcellular localization studies of the Isw2 protein.
Main Results:
- ISW2 is essential for early sporulation, with homozygous deletants arrested in G1 phase.
- Isw2 protein localizes to the cytoplasm in vegetative cells and accumulates in the nucleus during sporulation.
- Overexpression of ISW2 inhibits sporulation, while expression from a centromeric plasmid rescues the deletion phenotype.
Conclusions:
- The ISW2 gene plays a critical role in initiating meiosis and sporulation in yeast.
- Isw2 protein likely facilitates chromatin accessibility for transcriptional regulation of meiosis-specific genes.