Organization of the<i>GAL1–GAL10</i>intergeaic control region chromatin

Lohr, D.

doi:10.1093/nar/12.22.8457

Cited by 82 publications

(79 citation statements)

References 33 publications

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“…USA 83 (1986) within an -70-bp internucleosomal region as determined by nucleosome mapping experiments (14). Experiments by Lohr have shown the UAS region of the GALJ-GALJO promoter to be non-nucleosomal in nature (24,25).…”

Section: Discussionmentioning

confidence: 99%

Molecular analysis of the DNA sequences involved in the transcriptional regulation of the phosphate-repressible acid phosphatase gene (PHO5) of Saccharomyces cerevisiae.

Bergman¹,

McClinton²,

Madden³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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The expression of the PHOS gene of Saccharomyces cerevisiae is transcriptionally regulated in response to the level of inorganic phosphate present in the growth medium. We have identified, by DNA deletion analysis, the sequences (upstream activator sequences) that mediate this response. The sequence 5' CTGCACAATG 3' is present in two copies located within a 60-base-pair region. The presence of a single copy of the sequence is sufficient for the phosphatemediated transcriptional response. In addition, a DNA fragment that contains two copies of this sequence will act to repress transcription of a CYCI-lacZ fusion when placed either upstream or downstream of the CYCI activator sequence.The transcriptional regulation of genes in Saccharomyces cerevisiae is mediated through sequences located one hundred to several hundred bases upstream to the coding sequences (1)(2)(3)(4)(5) (12,13). Sequences at the 5' end determine a precise nucleosome positioning along the inactive gene sequence (14). In this study, using DNA deletions, we localize the sequences responsible for the transcriptional activation of PHOS transcription in response to the concentration of Pi in the growth medium. Also, using CYCI-PHOSlacZ gene fusions, we have demonstrated the presence of a "negative" factor involved in the repression of PHOS gene expression. MATERIALS AND METHODSStrains and Media. Strains of S. cerevisiae utilized in this study are listed in Table 1. Strains were grown in YCAD Fig. 1 for the structure of the wild-type plasmid p19UTT2) contains the E. coli plasmid pUC19, the yeast URA3 selectable marker, the 2-,um circle autonomously replicating segment, the CYCI transcription terminator, and a 1.2-kilobase Sal I/Pst I fragment containing the remainder of the PH05 gene. Thus, the plasmid contains unique BamHI and Sal I sites, allowing the analysis of deletions created within pBS-627 and its derivatives. pLG669Z, a CYCI-lacZ vector was obtained from L. Guarente (19). Derivatives either lacking the 430-bp Xho I fragment (p669-AXho) or with unique Xho I sites (p669-XP or p669-XD) were constructed in the laboratory.DNA Deletion Analysis and DNA Sequencing. DNA deletions within pBS-627 or its derivatives were created by either of two methods. (i) pBS-627 was cleaved with appropriate restriction enzymes, the single-stranded ends were filled-in with DNA polymerase I, and the plasmid was recircularized with T4 DNA ligase. (it) pBS-627 was cleaved with either TthlllI or Cla I and treated with BAL-31 for various time intervals. The purified DNA was subsequently treated with T4 DNA polymerase and recircularized with T4 DNA ligase in the presence of eitherXho I linker (for the TthlllI-cleaved sample) or Cla I linker (for the Cla I-cleaved sample). Individual E. coli transformants were screened by restriction analysis and subsequently the deletion fragment was purified by polyacrylamide gel electrophoresis. The fragment was Abbreviation: bp, base pair(s). 6070The publication costs of this article were defrayed in part by page charge payment...

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Section: Discussionmentioning

confidence: 99%

Molecular analysis of the DNA sequences involved in the transcriptional regulation of the phosphate-repressible acid phosphatase gene (PHO5) of Saccharomyces cerevisiae.

Bergman¹,

McClinton²,

Madden³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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“…First, one-fifth of digested DNA was separated by gel electrophoresis. Second, previously characterized GAL1 promoter sequences (3,8,18), one within a positioned nucleosome (GAL1 NB) and a second in an adjacent region (GAL1 NUB) that is rapidly digested by MNase, were amplified by qPCR from MNase-treated and untreated samples. The MNase concentration that resulted in mostly mononucleosomesized DNA with a GAL1 NUB/NB ratio of Ͻ15% was subjected to further qPCR using tiled SER3 primer pairs that amplify 38 unique SER3 sequences that range from 90 to 114 bp in size, with an average overlap of 69 bp between sequences.…”

Section: Methodsmentioning

confidence: 99%

Transcription Regulation by the Noncoding RNA SRG1 Requires Spt2-Dependent Chromatin Deposition in the Wake of RNA Polymerase II

Thebault

Boutin

Bhat

et al. 2011

Molecular and Cellular Biology

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Spt2 is a chromatin component with roles in transcription and posttranscriptional regulation. Recently, we found that Spt2 travels with RNA polymerase II (RNAP II), is involved in elongation, and plays important roles in chromatin modulations associated with this process. In this work, we dissect the function of Spt2 in the repression of SER3. This gene is repressed by a transcription interference mechanism involving the transcription of an adjacent intergenic region, SRG1, that leads to the production of a noncoding RNA (ncRNA). We find that Spt2 and Spt6 are required for the repression of SER3 by SRG1 transcription. Intriguingly, we demonstrate that these effects are not mediated through modulations of the SRG1 transcription rate. Instead, we show that the SRG1 region overlapping the SER3 promoter is occluded by randomly positioned nucleosomes that are deposited behind RNAP II transcribing SRG1 and that their deposition is dependent on the presence of Spt2. Our data indicate that Spt2 is required for the major chromatin deposition pathway that uses old histones to refold nucleosomes in the wake of RNAP II at the SRG1-SER3 locus. Altogether, these observations suggest a new mechanism of repression by ncRNA transcription involving a repressive nucleosomal structure produced by an Spt2-dependent pathway following RNAP II passage.

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“…The yeast GAL genes are repressed in glucose and highly induced in galactose, which is dependent upon the activator Gal4. In contrast to PHO5, the region to which Gal4 binds in the GAL1-10 locus-four sites spanning 135 bp-was originally believed to be nucleosome free (Lohr 1984(Lohr , 1993Fedor et al 1988;Fedor and Kornberg 1989;Cavalli and Thoma 1993). A recent study, however, showed that this region instead contains an unusual nucleosome, associated with the RSC (remodels the structure of chromatin) chromatin-remodeling complex, which protects a shorter fragment than canonical nucleosomes (Floer et al 2010).…”

Section: Histone Mutants Have Revealed New Facets About Transcriptionmentioning

confidence: 99%

Chromatin and Transcription in Yeast

2012

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Understanding the mechanisms by which chromatin structure controls eukaryotic transcription has been an intense area of investigation for the past 25 years. Many of the key discoveries that created the foundation for this field came from studies of Saccharomyces cerevisiae, including the discovery of the role of chromatin in transcriptional silencing, as well as the discovery of chromatin-remodeling factors and histone modification activities. Since that time, studies in yeast have continued to contribute in leading ways. This review article summarizes the large body of yeast studies in this field.

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Organization of theGAL1–GAL10intergeaic control region chromatin

Cited by 82 publications

References 33 publications

Molecular analysis of the DNA sequences involved in the transcriptional regulation of the phosphate-repressible acid phosphatase gene (PHO5) of Saccharomyces cerevisiae.

Molecular analysis of the DNA sequences involved in the transcriptional regulation of the phosphate-repressible acid phosphatase gene (PHO5) of Saccharomyces cerevisiae.

Transcription Regulation by the Noncoding RNA SRG1 Requires Spt2-Dependent Chromatin Deposition in the Wake of RNA Polymerase II

Chromatin and Transcription in Yeast

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