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Query: UNIPROT:P00492 (hypoxanthine-guanine phosphoribosyltransferase)
 2,385 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Recent studies suggest that enhancers may increase the accessibility of chromatin to transcription factors. To test the effects of a viral enhancer on chromatin accessibility, we have inserted minigenes with or without the polyomavirus enhancer into the third exon of the hypoxanthine phosphoribosyltransferase (HPRT) gene by homologous recombination and have prepared high-resolution maps of gene accessibility by using a novel polymerase chain reaction assay for DNase I sensitivity. In its native state, we find that the HPRT gene has low sensitivity to DNase I in fibrosarcoma cells. Insertion of the polyomavirus enhancer and neo reporter gene into exon 3 confers altered HPRT DNase I sensitivity for several kilobases on either side of the enhancer. The changes in DNase I sensitivity peak near the enhancer and decline with distance from the enhancer. The increase in HPRT DNase I sensitivity persisted when the tk promoter was deleted from the inserted construct but disappeared when the enhancer was deleted. These experiments identify the polyomavirus enhancer as a cis-acting initiator of chromatin accessibility.
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PMID:The polyomavirus enhancer activates chromatin accessibility on integration into the HPRT gene. 133 45

We investigated the conformation of the X-linked mouse hypoxanthine-guanine phosphoribosyltransferase gene (HPRT) promoter region both in chromatin from the active and inactive X chromosomes with DNase I and in naked supercoiled DNA with S1 nuclease. A direct comparison of the chromatin structures of the active and inactive mouse HPRT promoter regions was performed by simultaneous DNase I treatment of the active and inactive X chromosomes in the nucleus of interspecies hybrid cells from Mus musculus and Mus caroli. Using a restriction fragment length polymorphism to distinguish between the active and inactive HPRT promoters, we found a small but very distinct difference in the DNase I sensitivity of active versus inactive chromatin. We also observed a single DNase I-hypersensitive site in the immediate area of the promoter which was present only on the active X chromosome. Analysis of the promoter region by S1 nuclease digestion of supercoiled plasmid DNA showed an S1-sensitive site which maps adjacent to or within the DNase I-hypersensitive site found in chromatin but upstream of the region minimally required for normal HPRT gene expression.
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PMID:Nuclease sensitivity of the mouse HPRT gene promoter region: differential sensitivity on the active and inactive X chromosomes. 282 12

To examine the association between chromatin structure and gene expression at the human hypoxanthine phosphoribosyltransferase (HPRT) locus, DNase I sensitivity of active and inactive genes was analyzed. In a set of human-hamster hybrid lines containing either an active or an inactive human X chromosome, or a derivative of the latter in which the HPRT gene was reactivated by 5-azacytidine treatment, only the promoter region of the gene was found to contain a hypersensitive domain, and its presence was strictly correlated with gene activity. An S1 nuclease-sensitive site was mapped upstream from the DNase I hypersensitive domain using supercoiled plasmids. The overall level of DNase I sensitivity in the interior of the HPRT gene was also assessed by comparing the degradation of polymorphic restriction fragments on active and inactive alleles in both polyclonal and monoclonal lines of female human cells. In these internally controlled experiments, the active X chromosome was found to be approximately twofold more susceptible to DNase I digestion than the inactive X chromosome.
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PMID:Comparative study of DNase I sensitivity at the X-linked human HPRT locus. 283 22

DNA sequences of the X-chromosome-linked hypoxanthine phosphoribosyltransferase (HPRT) and glucose 6-phosphate dehydrogenase (G6PD) genes have revealed the presence of clusters of CpG dinucleotides, raising the possibility that such clusters are involved in the control of expression of these genes, which are expressed in all tissues. Although CpG clusters are not exclusive features of the X chromosome, the analysis of X-linked genes provides the means to determine whether CpG clusters are control elements; one of the two homologous X loci in female mammals is not expressed, so that active and inactive versions of the gene can be compared. In fact, it has been shown that these CpG clusters are undermethylated when the gene is active and extensively methylated when the gene is inactive. In addition to hypomethylation, chromatin hypersensitivity to endonuclease digestion is a known hallmark of regulatory sequences in eukaryotic genes. We report here that the CpG clusters of the active hprt and g6pd genes are not only undermethylated, but also hypersensitive to MspI, DNase I and S1 nuclease, further supporting the suggestion that they are involved in the control of expression of these genes.
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PMID:Clusters of CpG dinucleotides implicated by nuclease hypersensitivity as control elements of housekeeping genes. 298 78

To investigate the possible contribution of intragenic differentially methylated cytosines to X-linked gene expression, we examined DNA-protein interactions in a region in intron 3 of the human hypoxanthine phosphoribosyltransferase gene which contains at least one HpaII site methylated specifically on the active X. In vitro DNase I footprinting experiments using unmethylated DNA and HeLa nuclear extract identified three footprints (I-III). Footprints I and III flank an Alu repeat containing the HpaII site(s), one of which is contained within footprint II. Although methylation of the HpaII site had no effect on footprint II binding interactions, methylation of nearby CpGs substantially reduced the formation of three of the specific DNA-protein complexes binding to footprint II in mobility shift assays. Additionally, an A+T rich region immediately 5' to the HpaII-containing Alu repeat was found to bind specifically to nuclear matrices in vitro. We suggest that differential methylation of CpGs may affect the binding of regulatory proteins in vivo, and that interactions between the footprint proteins and those binding to the matrix attachment region may be involved in controlling X-linked Hprt expression.
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PMID:Intragenic matrix attachment and DNA-protein interactions in the human X-linked Hprt gene. 757 42

An intragenic region of the wallaroo (Macropus robustus) hypoxanthine phosphoribosyltransferase gene which contains three active X-specific methylated cytosines was examined for protein(s) binding. In vitro DNase I footprinting of unmethylated DNA identified three footprints, one of which (footprint I) contained two of the known differentially methylated sites (a HpaII and a HhaI site). Methylation of the footprint I HpaII site only, abolished the formation of several, specific DNA-protein complexes in mobility shift assays. UV cross-linking experiments indicated that polypeptides involved in the methylation sensitive interactions with footprint I had molecular weights ranging from 72 to 48 kDa. Analogous results were obtained with nuclear extracts from both eutherian and metatherian cells, indicating that these proteins are conserved. We suggest that the binding of these proteins to the inactive X may play some role in gene silencing, with the active gene being protected from this effect by methylation of the binding site.
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PMID:Methylation sensitive protein binding to an intragenic active X-specific methylated region in the M. robustus Hprt gene. 913 Oct 16

Differential chromatin structure is one of the hallmarks distinguishing active and inactive genes. For the X-linked human hypoxanthine phosphoribosyltransferase gene (HPRT), this difference in chromatin structure is evident in the differential general DNase I sensitivity and hypersensitivity of the promoter regions on active versus inactive X chromosomes. Here we characterize the nucleosomal organization responsible for the differential chromatin structure of the active and inactive HPRT promoters. The micrococcal nuclease digestion pattern of chromatin from the active allele in permeabilized cells reveals an ordered array of translationally positioned nucleosomes in the promoter region except over a 350-bp region that is either nucleosome free or contains structurally altered nucleosomes. This 350-bp region includes the entire minimal promoter and all of the multiple transcription initiation sites of the HPRT gene. It also encompasses all of the transcription factor binding sites identified by either dimethyl sulfate or DNase I in vivo footprinting of the active allele. In contrast, analysis of the inactive HPRT promoter reveals no hypersensitivity to either DNase I or a micrococcal nuclease and no translational positioning of nucleosomes. Although nucleosomes on the inactive promoter are not translationally positioned, high-resolution DNase I cleavage analysis of permeabilized cells indicates that nucleosomes are rotationally positioned over a region of at least 210 bp on the inactive promoter, which coincides with the 350-bp nuclease-hypersensitive region on the active allele, including the entire minimal promoter. This rotational positioning of nucleosomes is not observed on the active promoter. These results suggest a model in which the silencing of the HPRT promoter during X chromosome inactivation involves remodeling a transcriptionally competent, translationally positioned nucleosomal array into a transcriptionally repressed architecture consisting of rotationally but not translationally positioned nucleosomal arrays.
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PMID:Nucleosomes are translationally positioned on the active allele and rotationally positioned on the inactive allele of the HPRT promoter. 1160 4