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A novel mechanism for target gene-specific SWI/SNF recruitment via the Snf2p N-terminus.

Weider M, Schröder A, Klebl F, Sauer N - Nucleic Acids Res. (2011)

Bottom Line: Chromatin-remodeling complexes regulate the expression of genes in all eukaryotic genomes.Here, we show that the N-terminus of Snf2p, the chromatin remodeling core unit of the SWI/SNF complex, is essential for the expression of VHT1, the gene of the plasma membrane H(+)/biotin symporter, and of BIO5, the gene of a 7-keto-8-aminopelargonic acid transporter, biotin biosynthetic precursor. chromatin immunoprecipitation (ChIP) analyses demonstrate that Vhr1p, the transcriptional regulator of VHT1 and BIO5 expression, is responsible for the targeting of Snf2p to the VHT1 promoter at low biotin.We identified an Snf2p mutant, Snf2p-R(15)C, that specifically abolishes the induction of VHT1 and BIO5 but not of other Snf2p-regulated genes, such as GAL1, SUC2 or INO1.

View Article: PubMed Central - PubMed

Affiliation: Molekulare Pflanzenphysiologie, FAU Erlangen-Nürnberg, Staudtstraße 5, D-91058 Erlangen, Germany.

ABSTRACT
Chromatin-remodeling complexes regulate the expression of genes in all eukaryotic genomes. The SWI/SNF complex of Saccharomyces cerevisiae is recruited to its target promoters via interactions with selected transcription factors. Here, we show that the N-terminus of Snf2p, the chromatin remodeling core unit of the SWI/SNF complex, is essential for the expression of VHT1, the gene of the plasma membrane H(+)/biotin symporter, and of BIO5, the gene of a 7-keto-8-aminopelargonic acid transporter, biotin biosynthetic precursor. chromatin immunoprecipitation (ChIP) analyses demonstrate that Vhr1p, the transcriptional regulator of VHT1 and BIO5 expression, is responsible for the targeting of Snf2p to the VHT1 promoter at low biotin. We identified an Snf2p mutant, Snf2p-R(15)C, that specifically abolishes the induction of VHT1 and BIO5 but not of other Snf2p-regulated genes, such as GAL1, SUC2 or INO1. We present a novel mechanism of target gene-specific SWI/SNF recruitment via Vhr1p and a conserved N-terminal Snf2p domain.

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Growth test to assay phenotypes of ySWI/SNF mutants. Yeast strains were grown on the indicated media to assay the following phenotypes: slow growth (YPD), inability to use the sugars galactose or raffinose as carbon sources (YPGal and YPRaf), and inositol auxotrophy (SD w/o inositol). The SNF2 WT (top) and the Δsnf2 mutant (bottom) were used as controls to assay the growth behavior of the snf2-R15C mutant. In addition, the effects of SNF2 expression from one of the complementing multicopy plasmids (snf2-R15C + SNF2mc; Figure 1A) or from an SNF2-containing single copy plasmid (snf2-R15C + SNF2sc) were analyzed. To allow growth on the same medium, control strains were transformed with the empty plasmid YEp24. Cells were spotted on plates after serial 1:8 dilutions.
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Figure 6: Growth test to assay phenotypes of ySWI/SNF mutants. Yeast strains were grown on the indicated media to assay the following phenotypes: slow growth (YPD), inability to use the sugars galactose or raffinose as carbon sources (YPGal and YPRaf), and inositol auxotrophy (SD w/o inositol). The SNF2 WT (top) and the Δsnf2 mutant (bottom) were used as controls to assay the growth behavior of the snf2-R15C mutant. In addition, the effects of SNF2 expression from one of the complementing multicopy plasmids (snf2-R15C + SNF2mc; Figure 1A) or from an SNF2-containing single copy plasmid (snf2-R15C + SNF2sc) were analyzed. To allow growth on the same medium, control strains were transformed with the empty plasmid YEp24. Cells were spotted on plates after serial 1:8 dilutions.

Mentions: Compared with WT cells, Δsnf2 mutants exhibit slow growth on glucose [Figure 5 and (42,43)], fail to grow on galactose and raffinose (42,43), and are auxotrophs for inositol (44). This was re-confirmed in comparative growth analyses on media with galactose (YPGal) or raffinose (YPRaf) as sole carbon source, on medium w/o inositol (SD w/o inositol), or on glucose-containing yeast peptone dextrose (YPD) medium (Figure 6). Surprisingly, however, the snf2-R15C mutant did not show any of these phenotypes and behaved like the WT on all media (Figure 6). Moreover, this growth could not be further improved by expressing WT SNF2 from one of the multicopy plasmids (snf2-R15C + SNF2mc) identified in the complementation screening (Figure 1A) or from the single copy plasmid (snf2-R15C + SNF2sc) used to express SNF2 in the AMYmut153 strain (Figure 1A). This demonstrated that the snf2-R15C mutation does not exhibit any of the previously published growth phenotypes of a Δsnf2 deletion mutant. It rather suggested that the snf2-R15C mutation is hypomorphic allele that specifically affects the regulation by Vhr1p and the targeting of ySWI/SNF to the VHRE.Figure 6.


A novel mechanism for target gene-specific SWI/SNF recruitment via the Snf2p N-terminus.

Weider M, Schröder A, Klebl F, Sauer N - Nucleic Acids Res. (2011)

Growth test to assay phenotypes of ySWI/SNF mutants. Yeast strains were grown on the indicated media to assay the following phenotypes: slow growth (YPD), inability to use the sugars galactose or raffinose as carbon sources (YPGal and YPRaf), and inositol auxotrophy (SD w/o inositol). The SNF2 WT (top) and the Δsnf2 mutant (bottom) were used as controls to assay the growth behavior of the snf2-R15C mutant. In addition, the effects of SNF2 expression from one of the complementing multicopy plasmids (snf2-R15C + SNF2mc; Figure 1A) or from an SNF2-containing single copy plasmid (snf2-R15C + SNF2sc) were analyzed. To allow growth on the same medium, control strains were transformed with the empty plasmid YEp24. Cells were spotted on plates after serial 1:8 dilutions.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3105400&req=5

Figure 6: Growth test to assay phenotypes of ySWI/SNF mutants. Yeast strains were grown on the indicated media to assay the following phenotypes: slow growth (YPD), inability to use the sugars galactose or raffinose as carbon sources (YPGal and YPRaf), and inositol auxotrophy (SD w/o inositol). The SNF2 WT (top) and the Δsnf2 mutant (bottom) were used as controls to assay the growth behavior of the snf2-R15C mutant. In addition, the effects of SNF2 expression from one of the complementing multicopy plasmids (snf2-R15C + SNF2mc; Figure 1A) or from an SNF2-containing single copy plasmid (snf2-R15C + SNF2sc) were analyzed. To allow growth on the same medium, control strains were transformed with the empty plasmid YEp24. Cells were spotted on plates after serial 1:8 dilutions.
Mentions: Compared with WT cells, Δsnf2 mutants exhibit slow growth on glucose [Figure 5 and (42,43)], fail to grow on galactose and raffinose (42,43), and are auxotrophs for inositol (44). This was re-confirmed in comparative growth analyses on media with galactose (YPGal) or raffinose (YPRaf) as sole carbon source, on medium w/o inositol (SD w/o inositol), or on glucose-containing yeast peptone dextrose (YPD) medium (Figure 6). Surprisingly, however, the snf2-R15C mutant did not show any of these phenotypes and behaved like the WT on all media (Figure 6). Moreover, this growth could not be further improved by expressing WT SNF2 from one of the multicopy plasmids (snf2-R15C + SNF2mc) identified in the complementation screening (Figure 1A) or from the single copy plasmid (snf2-R15C + SNF2sc) used to express SNF2 in the AMYmut153 strain (Figure 1A). This demonstrated that the snf2-R15C mutation does not exhibit any of the previously published growth phenotypes of a Δsnf2 deletion mutant. It rather suggested that the snf2-R15C mutation is hypomorphic allele that specifically affects the regulation by Vhr1p and the targeting of ySWI/SNF to the VHRE.Figure 6.

Bottom Line: Chromatin-remodeling complexes regulate the expression of genes in all eukaryotic genomes.Here, we show that the N-terminus of Snf2p, the chromatin remodeling core unit of the SWI/SNF complex, is essential for the expression of VHT1, the gene of the plasma membrane H(+)/biotin symporter, and of BIO5, the gene of a 7-keto-8-aminopelargonic acid transporter, biotin biosynthetic precursor. chromatin immunoprecipitation (ChIP) analyses demonstrate that Vhr1p, the transcriptional regulator of VHT1 and BIO5 expression, is responsible for the targeting of Snf2p to the VHT1 promoter at low biotin.We identified an Snf2p mutant, Snf2p-R(15)C, that specifically abolishes the induction of VHT1 and BIO5 but not of other Snf2p-regulated genes, such as GAL1, SUC2 or INO1.

View Article: PubMed Central - PubMed

Affiliation: Molekulare Pflanzenphysiologie, FAU Erlangen-Nürnberg, Staudtstraße 5, D-91058 Erlangen, Germany.

ABSTRACT
Chromatin-remodeling complexes regulate the expression of genes in all eukaryotic genomes. The SWI/SNF complex of Saccharomyces cerevisiae is recruited to its target promoters via interactions with selected transcription factors. Here, we show that the N-terminus of Snf2p, the chromatin remodeling core unit of the SWI/SNF complex, is essential for the expression of VHT1, the gene of the plasma membrane H(+)/biotin symporter, and of BIO5, the gene of a 7-keto-8-aminopelargonic acid transporter, biotin biosynthetic precursor. chromatin immunoprecipitation (ChIP) analyses demonstrate that Vhr1p, the transcriptional regulator of VHT1 and BIO5 expression, is responsible for the targeting of Snf2p to the VHT1 promoter at low biotin. We identified an Snf2p mutant, Snf2p-R(15)C, that specifically abolishes the induction of VHT1 and BIO5 but not of other Snf2p-regulated genes, such as GAL1, SUC2 or INO1. We present a novel mechanism of target gene-specific SWI/SNF recruitment via Vhr1p and a conserved N-terminal Snf2p domain.

Show MeSH
Related in: MedlinePlus