Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.31.1 (micrococcal nuclease)
 2,818 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The beta-like globin genes require the upstream locus control region (LCR) for proper expression. The active elements of the LCR coincide with strong erythroid-specific DNase I-hypersensitive sites (HSs). We have used 5' HS4 as a model to study the formation of these HSs. Previously, we identified a 101 bp element that is required for the formation of this HS. This element binds six proteins in vitro. We now report a mutational analysis of the HS4 HS-forming element (HSFE). This analysis indicates that binding sites for the hematopoietic transcription factors NF-E2 and GATA-1 are required for the formation of the characteristic chromatin structure of the HS following stable transfection into murine erythroleukemia cells. Similarly arranged NF-E2 and GATA binding sites are present in the other HSs of the human LCR, as well as in the homologous mouse and goat sequences and the chicken beta-globin enhancer. A combination of DNase I and micrococcal nuclease sensitivity assays indicates that the characteristic erythroid-specific hypersensitivity of HS4 to DNase I is the result of tissue-specific alterations in both nucleosome positioning and tertiary DNA structure.
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PMID:NF-E2 and GATA binding motifs are required for the formation of DNase I hypersensitive site 4 of the human beta-globin locus control region. 782 82

Thromboxane A(2), a potent mediator of vasoconstriction and platelet aggregation, is synthesized from prostaglandin H(2) by thromboxane synthase (TXAS). We report here on promoter analyses of human TXAS using in vitro transcription and in vivo transfection methods. The 39-bp core promoter, containing both TATA and initiator elements, accurately initiates transcription in an orientation-dependent manner in a cell-free transcription system. Mutation of either TATA or initiator abolished transcriptional activity, but the upstream sequence had no effect on TXAS promoter activities in vitro, suggesting that the core promoter is sufficient for transcriptional activity from a naked DNA template. In contrast, mutation of both elements drastically decreased the promoter activity in vivo. Furthermore, TXAS proximal promoter containing the nucleotides -90 to -56 relative to the transcriptional start site was necessary for promoter transactivation in vivo. Transcriptional activities from 5'-deletion mutants indicated that the effects of the nucleotides -90/-56 were more pronounced in stably transfected cells (a 150-fold difference) than in the transiently transfected cells (an 8-fold difference), reflecting the effects of TXAS transcriptional activation from replicating and nonreplicating DNA templates. Partial micrococcal nuclease digestion indicated that the sequence -90/-56, where the NF-E2 site is located, is associated with alterations of nucleosomal structure at the regions of promoter and reporter gene but not the region further downstream. Mutagenesis and forced expression studies demonstrated a critical role of p45 NF-E2 in controlling TXAS expression under native chromatin conditions. Band shifting and chromatin immunoprecipitation assays indicated that p45 NF-E2 was bound to the TXAS promoter in vitro and in vivo. Indirect end labeling and ligation-mediated PCR analyses further demonstrated that the occupation of TXAS promoter NF-E2 site was associated with disruption of nucleosomal structure. Collectively, these results indicate that binding of NF-E2 is critical both for alteration of the nucleosomal structure and for activation of the TXAS promoter, whereas the TATA and initiator elements are essential for transcription.
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PMID:Transcriptional control of the human thromboxane synthase gene in vivo and in vitro. 1195 85

When transcription is initiated under repressive conditions, such as when chromatin are packed together, binding followed by the functioning of key components in the transcriptional apparatus should be appropriately facilitated in the chromatin architecture. We provide evidence that the erythroid-specific enhancer- binding protein NF-E2 interacts with the cognate motif at DNase I-hypersensitive site 2 of the human beta-globin locus control region in a repressive state. The nucleosome containing the NF-E2-binding site showed characteristic rotational and translational phases in vitro. The binding site had less affinity to the histone octamers than nearby regions while showing greater accessibility to DNase I and micrococcal nuclease. Furthermore, the motif was recognized by the exogenous NF-E2 protein expressed in HeLa cells, which have a repressive state of chromatin at the beta-globin locus, as shown by ligation-mediated PCR and chromatin immunoprecipitation assay. These lines of evidence indicate that NF-E2 interacts with the cognate motif on the nucleosome before chromatin is remodeled.
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PMID:Interaction of NF-E2 in the human beta-globin locus control region before chromatin remodeling. 1250 25