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. 2000 Aug;20(16):5960-73.
doi: 10.1128/MCB.20.16.5960-5973.2000.

Genetic interactions between TFIIS and the Swi-Snf chromatin-remodeling complex

J K Davie  1 , C M Kane
Affiliations

Genetic interactions between TFIIS and the Swi-Snf chromatin-remodeling complex

J K Davie et al. Mol Cell Biol. 2000 Aug.

Abstract

The eukaryotic transcript elongation factor TFIIS enables RNA polymerase II to read through blocks to elongation in vitro and interacts genetically with a variety of components of the transcription machinery in vivo. In Saccharomyces cerevisiae, the gene encoding TFIIS (PPR2) is not essential, and disruption strains exhibit only mild phenotypes and an increased sensitivity to 6-azauracil. The nonessential nature of TFIIS encouraged the use of a synthetic lethal screen to elucidate the in vivo roles of TFIIS as well as provide more information on other factors involved in the regulation of transcript elongation. Several genes were identified that are necessary for either cell survival or robust growth when the gene encoding TFIIS has been disrupted. These include UBP3, KEX2, STT4, and SWI2/SNF2. SWI1 and SNF5 disruptions were also synthetically lethal with ppr2Delta, suggesting that the reduced ability to remodel chromatin confers the synthetic phenotype. The synthetic phenotypes show marked osmosensitivity and cytoskeletal defects, including a terminal hyperelongated bud phenotype with the Swi-Snf complex. These results suggest that genes important in osmoregulation, cell membrane synthesis and integrity, and cell division may require the Swi-Snf complex and TFIIS for efficient transcription. The detection of these genetic interactions provides another functional link between the Swi-Snf complex and the elongation machinery.

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Figures

FIG. 1
FIG. 1
Sensitive growth of synthetic lethal mutant 28A. (A) The synthetic lethal mutant 28A, a wild-type strain (CH1305), and mutant 28A transformed with the SNF2-containing library plasmid pLPSNF2 were streaked onto medium with inositol added. (B) The three strains were streaked onto medium without inositol. The plates were photographed after 4 days of growth at 30°C. (C) Wild-type (CH1305), mutant 28A, and mutant 28A transformed with the SNF2-containing library plasmid pLPSNF2 were streaked on dextrose medium. The plate was photographed after 4 days of anaerobic growth at 30°C. (D) The same strains were streaked onto raffinose medium and grown anaerobically for 4 days at 30°C prior to photography.
FIG. 2
FIG. 2
A snf2Δ allele is synthetically lethal in combination with a ppr2Δ allele. (A) Tetrad analysis following sporulation of an snf2Δ::kanr/SNF2 ppr2Δ::URA3/PPR2 diploid (CMKy38). The tetrads were dissected on YPD medium, and the YPD plate was photographed after 7 days of growth at 30°C. (B) The snf2Δ::kanr spores were identified by their G418r phenotype, the ppr2Δ::URA3 spores were identified by their Ura+ phenotype, and TRP1 spores were identified by their Trp+ phenotype. The positions of inviable spores are shaded gray, the box containing the genotype deduced from the other segregants. The presence of the TRP1+ gene and the trp1 mutation indicated with a T and with a t, respectively.
FIG. 3
FIG. 3
Hyperelongated bud phenotype. The snf2Δ/SNF2 ppr2Δ/ppr2Δ diploid (CMKy46) under 100× magnification. Cells were visualized by differential interference contrast microscopy (see Materials and Methods). Approximately 45% of cells had this morphology.
FIG. 4
FIG. 4
A snf2Δ allele is synthetically lethal with a ppr2Δ allele in two unrelated genetic backgrounds. (A) Tetrad analysis following sporulation of a snf2Δ::kanr/SNF2 ppr2Δ::URA3/PPR2 diploid derived from YPH499/500 (CMKy77). The tetrads were dissected on YPD medium, and the YPD plate was photographed after 7 days of growth at 30°C. (B) The snf2Δ::kanr spores were identified by their G418r phenotype, and ppr2Δ::URA3 spores were identified by their Ura+ phenotype. Inviable spore positions are marked with a shaded box containing the genotype deduced from the other segregants. (C) Tetrad analysis following sporulation of an snf2Δ::kanr/SNF2 ppr2Δ::URA3/PPR2 diploid derived from W303 (CMKy79). The tetrads were dissected on YPD medium, and the YPD plate was photographed after 7 days of growth at 30°C. (D) The snf2Δ::kanr spores were identified by their G418r phenotype, and ppr2Δ::URA3 spores were identified by their Ura+ phenotype. Inviable spore positions are marked with a shaded box containing the genotype deduced from the other segregants.
FIG. 5
FIG. 5
Both swi1Δ and snf5Δ alleles are synthetically lethal in combination with a ppr2Δ allele. (A) Tetrad analysis following sporulation of an swi1Δ::kanr/SWI1 ppr2Δ::URA3/PPR2 diploid (CMKy60). The tetrads were dissected on YPD medium, and the YPD plate was photographed after 7 days of growth at 30°C. (B) The swi1Δ::kanr spores were identified by their G418r phenotype, the ppr2Δ::URA3 spores were identified by their Ura+ phenotype, and TRP1 spores were identified by their Trp+ phenotype. Inviable spore positions are marked with a shaded box containing the genotype deduced from the other segregants. The presence of the TRP1+ gene is indicated with a T, and that of the trp1 mutation is indicated with a t. (C) Tetrad analysis following sporulation of an snf5Δ::kanr/SNF5 ppr2Δ::URA3/PPR2 diploid (CMKy64). The tetrads were dissected on YPD medium, and the YPD plate was photographed after 8 days of growth at 30°C. (D) The snf5Δ::kanr spores were identified by their G418r phenotype, the ppr2Δ::URA3 spores were identified by their Ura+ phenotype, and TRP1 spores were identified by their Trp+ phenotype. Inviable spore positions are marked with a shaded box containing the genotype deduced from the other segregants. The presence of the TRP1+ gene is indicated with a T, and that of the trp1 allele is indicated with a t.
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