5-Azadeoxycytidine-induced Chromatin Remodeling of the Inactive X-linked HPRT Gene Promoter Occurs prior to Transcription Factor Binding and Gene Reactivation

Litt, Michael D.; Hansen, R. Scott; Hornstra, Ian; Gartler, Stanley M.; Yang, Thomas P.

doi:10.1074/jbc.272.23.14921

Cited by 22 publications

(28 citation statements)

References 39 publications

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“…Because differential DNA methylation of the active and inactive HPRT promoters is a prominent feature of the HPRT locus and because demethylation of the promoter by 5aCdr is closely associated with the remodeling of chromatin in the promoter region (27,44), it was conceivable that DNA methylation could play a role in establishing and/or maintaining the differential nucleosomal architecture of the active and inactive HPRT promoter regions. However, our analysis of in vitro chromatin assembly on methylated and unmethylated HPRT promoter templates (Fig.…”

Section: Discussionmentioning

confidence: 99%

“…Litt et al (27) have shown that, during the course of HPRT gene reactivation on the inactive X chromosome by 5aCdr, chromatin remodeling of the promoter region from a nuclease-resistant to a nuclease-accessible conformation occurs prior to both transcription factor binding and the appearance of mRNA. Presumably, the 5aCdr-induced remodeling of the promoter region consists of converting the nucleosomal organization on the inactive promoter, where nucleosomes are not translationally positioned across the promoter region but rather are rotationally positioned over key regulatory regions, to a transcriptionally competent structure that contains translationally positioned nucleosomal arrays flanking a nucleosome-free region.…”

Section: Discussionmentioning

confidence: 99%

“…8) suggests that the protein bound to the AP-2 site may act as a boundary element to maintain the positioning of the translationally positioned nucleosome array upstream of the AP-2 site. However, Litt et al (27) have shown that, during the reactivation of the HPRT gene by 5aCdr, chromatin remodeling from a nuclease-resistant to a nuclease-accessible conformation occurs prior to the detectable binding of transcription factors to the promoter region (see below). Even if the protein bound to the AP-2 site does not mediate the initial remodeling and opening of chromatin during 5aCdr-induced reactivation, it may nevertheless be involved in positioning the first nucleosome of the upstream array and preventing nucleosomes from assembling on or sliding into the highly nucleaseaccessible 350-bp region in the active allele.…”

Section: Discussionmentioning

confidence: 99%

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Nucleosomes Are Translationally Positioned on the Active Allele and Rotationally Positioned on the Inactive Allele of the HPRT Promoter

Chen

Yang

2001

Molecular and Cellular Biology

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Differential chromatin structure and accessibility, particularly at the promoter, have long been recognized as characteristics that distinguish active from inactive genes. Active genes are in general more accessible to regulatory factors than inactive genes, as indicated by nuclease sensitivity. In addition, the promoters of active genes often exhibit marked DNase I hypersensitivity, especially in the vicinity of transcription factor binding sites (10,20). This hypersensitivity is postulated to be due to changes in the chromatin architecture of the promoter and may represent nucleosomal remodeling or displacement, stretches of single-stranded DNA, torsionally stressed DNA, or other distortions in chromatin structure arising from factor binding (18,20,62).The functional effect of nucleosomes on transcription initiation is thought to be repressive since in vitro assembly of nucleosomal arrays on DNA templates drastically reduces the capacity of these templates to support basal transcription (25,35,36,57). Furthermore, the differential accessibility and transcriptional potential of chromatin structure in active versus inactive promoters are often associated with differential nucleosomal organization (3,5,23,30,46,62). Thus, remodeling the nucleosomal architecture of a promoter is likely to be an integral feature of mechanisms of gene activation and/or silencing, which may involve histone acetylation and chromatinremodeling complexes such as SWI/SNF (15,20,60).The organization of genomic DNA into nucleosomal arrays is defined by both the translational position of the nucleosome relative to the linear nucleotide sequence and the rotational orientation of the DNA helix relative to the surface of the histone octamer. The translational position of nucleosomes on a DNA template (i.e., the linear position of the nucleosome relative to the DNA sequence [46]) has been shown to affect the accessibility of cis-acting elements in the promoter to various transcription factors as well as the basal transcription complex. Whether a transcription factor binding site is incorporated into a nucleosomal core or is in the linker region between nucleosomes can dramatically affect its accessibility to its cognate binding protein(s) in vitro (25,58). However, the extent to which incorporating a transcription factor binding site into a nucleosomal core reduces the accessibility of that site can vary widely among transcription factors (4, 25). The rotational orientation of the DNA helix wound around a nucleosomal core also affects the accessibility of that DNA to transcription factors (25). For instance, the rotational orientation of the TATA box, the glucocorticoid response element, and the thyroid response element within a nucleosome strongly affects the accessibility of these elements to their cognate binding factors (14,24,59). These findings suggest that both the translational position and rotational orientation of cis-acting

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Nucleosomes Are Translationally Positioned on the Active Allele and Rotationally Positioned on the Inactive Allele of the HPRT Promoter

Chen

Yang

2001

Molecular and Cellular Biology

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“…For a number of genes, formation of promoter DNase I HS sites precedes active transcription [50][51][52], perhaps reflecting the activity of chromatin remodeling complexes. Indeed, maintenance of expression from the MoMuLV LTR requires Bramha, the catalytic subunit of one of the mammalian SWI/SNF complexes [53].…”

Section: Discussionmentioning

confidence: 99%

An Unmethylated 3' Promoter-proximal Region is Required for Efficient Transcription Initiation

Appanah¹,

Dickerson²,

Goyal³

et al. 2005

PLoS Genet

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The promoter regions of approximately 40% of genes in the human genome are embedded in CpG islands, CpG-rich regions that frequently extend on the order of one kb 39 of the transcription start site (TSS) region. CpGs 39 of the TSS of actively transcribed CpG island promoters typically remain methylation-free, indicating that maintaining promoterproximal CpGs in an unmethylated state may be important for efficient transcription. Here we utilize recombinasemediated cassette exchange to introduce a Moloney Murine Leukemia Virus (MoMuLV)-based reporter, in vitro methylated 1 kb downstream of the TSS, into a defined genomic site. In a subset of clones, methylation spreads to within ;320 bp of the TSS, yielding a dramatic decrease in transcript level, even though the promoter/TSS region remains unmethylated. Chromatin immunoprecipitation analyses reveal that such promoter-proximal methylation results in loss of RNA polymerase II and TATA-box-binding protein (TBP) binding in the promoter region, suggesting that repression occurs at the level of transcription initiation. While DNA methylation-dependent trimethylation of H3 lysine (K)9 is confined to the intragenic methylated region, the promoter and downstream regions are hypo-acetylated on H3K9/K14. Furthermore, DNase I hypersensitivity and methylase-based single promoter analysis (M-SPA) experiments reveal that a nucleosome is positioned over the unmethylated TATA-box in these clones, indicating that dense DNA methylation downstream of the promoter region is sufficient to alter the chromatin structure of an unmethylated promoter. Based on these observations, we propose that a DNA methylation-free region extending several hundred bases downstream of the TSS may be a prerequisite for efficient transcription initiation. This model provides a biochemical explanation for the typical positioning of TSSs well upstream of the 39 end of the CpG islands in which they are embedded.Citation: Appanah R, Dickerson DR, Goyal P, Groudine M, Lorincz MC (2007) An unmethylated 39 promoter-proximal region is required for efficient transcription initiation. PLoS Genet 3(2): e27.

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“…In contrast, the hypoxanthine phosphoribosyltransferase gene on the inactive X chromosome is not transcribed, and its promoter is occupied by nucleosomes, and it is not DNase I hypersensitive. Reactivation of the inactive allele by azadeoxycytidine is accompanied by the appearance of hypoxanthine phosphoribosyltransferase mRNA and the DHS (54). Similarly, ␤-globin transcription is dependent upon the erythroid Krueppel-like factor (EKLF) transcription factor.…”

Section: Significance Of Dhss At P53-regulated Promoters Under Nonindmentioning

confidence: 99%

Constitutive DNase I Hypersensitivity of p53-Regulated Promoters

Braastad

Han

Hendrickson

2003

Journal of Biological Chemistry

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The ability of p53 to alter, at the transcriptional level, the gene expression of downstream targets is critical for its role as a tumor suppressor. Most models of p53 activation postulate the stepwise recruitment by p53 of coactivators, histone acetyltransferases, and/or chromatin remodeling factors to a promoter region to facilitate the subsequent access of the general transcriptional machinery required for transcriptional induction. We demonstrate here, however, that the promoter regions for the p53 target genes, p21, 14-3-3, and KARP-1, exist in a constitutively open conformation that is readily accessible to DNase I. This conformation was not altered by DNA damage or by whether p53 was present or absent in the cell. In contrast, p53 response elements, which resided outside the immediate promoter regions, existed within DNase I-resistant chromatin domains. Thus, p53 activation of downstream target genes occurs without p53 inducing chromatin alterations detectable by DNase I accessibility at either the promoter or the response element. As such, these data support models of p53 activation that do not require extensive chromatin alterations to support cognate gene expression.The differential chromatin structure and nuclease accessibility at a promoter region has long been recognized as a key distinguishing feature between active and inactive genes (reviewed in Ref. 1). Almost invariably, the promoter regions of active genes are marked experimentally by hypersensitivity to restriction endonucleases (2, 3) or, more commonly, DNase I (4). Although this hypersensitivity can be caused by torsional or topological stress in the DNA and distortions in the chromatin structure resulting from transcription factor binding, it is generally caused by the absence of nucleosomes or their remodeling (1, 4). The paucity of nucleosomes, in turn, is a direct consequence of the repressive effect that nucleosomes have on the transcriptional machinery and the requirement to alleviate that effect for productive transcription to occur (5-7). Thus, inactive genes usually have promoters that are nucleosomal, are insensitive to DNase I, and are regarded as being in a closed confirmation whereas active genes usually have core promoters that are nucleosome-free, hypersensitive to DNase I, and in an open configuration. Consequently, one of the hallmarks of gene induction mechanisms is the remodeling of the nucleosomal architecture of a promoter as it is activated (5, 6, 8).Enormous strides have been made in the last decade in understanding transcriptional activation at the chromatin level. In particular, the activation of expression of many, although not all (9 -11), inducible eukaryotic genes is consistent with what can collectively be called recruitment models of gene activation (3, 7, 8) (reviewed in Ref. 12). These models usually require, in response to some extracellular cue, the induction of a specific transcription factor and the subsequent interaction of that factor with its cognate response element, which is invariably a cis-acting ...

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5-Azadeoxycytidine-induced Chromatin Remodeling of the Inactive X-linked HPRT Gene Promoter Occurs prior to Transcription Factor Binding and Gene Reactivation

Cited by 22 publications

References 39 publications

Nucleosomes Are Translationally Positioned on the Active Allele and Rotationally Positioned on the Inactive Allele of the HPRT Promoter

Nucleosomes Are Translationally Positioned on the Active Allele and Rotationally Positioned on the Inactive Allele of the HPRT Promoter

An Unmethylated 3' Promoter-proximal Region is Required for Efficient Transcription Initiation

Constitutive DNase I Hypersensitivity of p53-Regulated Promoters

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