Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.30.1 (S1 nuclease)
3,660 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chromosomal rearrangements involving the c-myc oncogene are a prevalent feature of plasmacytomas that arise after inoculating BALB/c mice with pristane and Abelson murine leukaemia virus (A-MuLV). With this observation in mind, we decided to determine if any genetic alterations of the c-myc locus could be observed in cells of a different type, when transformed in vitro by A-MuLV. Here we have analysed three independent A-MuLV-transformed NIH 3T3 lines (ANN-I, 54c12 and N25), and found that the c-myc locus is amplified 8-19-fold in each transformant. Quantitative S1 nuclease mapping performed on ANN-I and 54c12 RNAs demonstrated that: (1) c-myc messenger RNAs accumulated to double the levels found in NIH 3T3 cells; and (2) a shift in the use of the two normal c-myc transcription initiation sites (P1 and P2) occurred in favour of the 3' site, P2. Analysis of c-myc chromatin by DNase I treatment of 54c12 nuclei revealed that most, if not all, of the c-myc gene copies were transcriptionally competent. We present alternative ideas to explain why amplification of the c-myc gene occurs repeatedly in A-MuLV-transformed fibroblasts. Finally, we discuss our results in relation to the hypothesis linking the phenomenon of tumour progression with the amplification of oncogenes.
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PMID:Amplification and altered expression of the c-myc oncogene in A-MuLV-transformed fibroblasts. 299 29

By use of synchronized human HeLa S3 cells, a site sensitive to both DNase I and nuclease S1 was identified 50-150 base pairs upstream of the ATG codon of a cell cycle-dependent histone H4 gene. This site expanded to include a broad region of approximately equal to 300 base pairs sensitive to DNase I throughout S phase and then narrowed again to the original site after the completion of DNA replication. The level of nuclease S1 sensitivity was greatest during early S phase, when the gene is replicated and its transcription rate is maximal. The chromatin structure of the human beta-globin gene, which is not expressed in HeLa cells, was also analyzed throughout the cell cycle, and in no case was a sub-band seen as a result of DNase I or nuclease S1 digestion, nor were there any changes in nuclease sensitivity correlated with its replication. Thus the cell cycle-dependent chromatin alterations in this histone H4 gene appear to be due to the coupled replication and expression of this gene rather than simply its replication. These results suggest that histone genes, as compared with developmentally regulated genes, exhibit an "intermediate" level of regulation whereby the gene is never in a completely inactive conformation, but changes in chromatin structure occur as a function of the cell cycle and expression.
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PMID:A human histone H4 gene exhibits cell cycle-dependent changes in chromatin structure that correlate with its expression. 299 68

Using S1 nuclease protection assays, we have examined the representation of cell cycle-dependent H4 histone RNAs in the nuclear matrix and nonmatrix nuclear fractions of human cells. Cytoplasmic and nuclear fractions were prepared from exponentially growing HeLa S3 cells by double detergent (sodium deoxycholate and NP40) lysis. The nuclear matrix and nonmatrix nuclear fractions were then prepared by digestion of nuclei with RNase-free DNase I and subsequent high-salt [0.4 M (NH4)2SO4] extraction. Subcellular fractions were characterized by 1) DNA, RNA, and protein composition; 2) electrophoretic analysis of the proteins in each fraction; 3) the representation of 45S ribosomal RNA precursors and processed 18S and 28S ribosomal RNAs; and 4) the presence of mitochondrial RNAs. In contrast to ribosomal and messenger RNA precursors, which are largely associated with the nuclear matrix, the human H4 histone RNAs in the nucleus were found predominantly in the nonmatrix nuclear fraction. The presence of H4 histone RNA in the nonmatrix nuclear fraction appeared to be coupled to DNA replication, since inhibition of DNA synthesis by hydroxyurea resulted in a loss of histone RNA from the nucleus. Our results suggest either that the association of histone RNAs with the nuclear matrix is very transient or that posttranscriptional modifications of the rapidly processed histone gene transcripts do not involve the nuclear matrix.
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PMID:Localization of human histone gene transcripts predominantly in the nonmatrix nuclear fraction. 301 90

We have analyzed the chromatin structure of a region that encompasses 14.4 X 10(3) base-pairs of the chicken histone H5 locus in adult erythroid cells at different stages of maturation. Seven of eight major lineage-specific DNase I-hypersensitive sites, some of which show complex substructure, were found in the flanking regions of the gene. The hypersensitivity of some of these sites is modulated during erythrocyte maturation in a way that parallels the transcriptional activity of the gene. DNase I, micrococcal nuclease, and S1 nuclease recognize the same regions, which differ from those cleaved by S1 on supercoiled plasmid DNA. This suggests that hypersensitivity of DNA in chromatin reflects a greater accessibility of the DNA rather than its altered conformation. The DNA sequence of some of the DNase I target sites contains repeated motifs, (T-C-C-C)2, (T-C-C)2, (T-G-G-G-G)2, which are found in the hypersensitive sites of other genes. Detailed analysis across sections of the H5 gene and flanking sequences revealed differences in the DNase I sensitivity of the different regions examined. Notably, the first one-third of the gene is more sensitive than the rest. The sequences downstream from the region where most RNA polymerases terminate transcription were found to be the most resistant.
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PMID:Fine analysis of the active H5 gene chromatin of chicken erythroid cells at different stages of differentiation. 302 21

Expression of the genes in the phosphate regulon, including the pstS (phoS) and phoB genes, is positively regulated by PhoB protein when phosphate is limited. We purified PhoB protein from overproducing cells and studied its interaction with the pstS gene. It binds specifically to the DNA fragment containing the promoter region of pstS. The transcription initiation site of the gene in vivo was identified by S1 nuclease mapping and primer-extension experiments. In-vitro transcription of pstS was activated by the PhoB protein, and the initiation site of transcription agreed with the in-vivo initiation site. Activation of in-vitro transcription by PhoB protein required both the normal sigma factor (sigma 70) and core RNA polymerase. PhoB protein binding sites on the promoter regions of pstS and phoB were determined by footprinting experiments with DNase I and a methylating agent. In both cases the protein binds to the pho box, the concensus sequence shared by regulatory regions of genes in the phosphate regulon. Our findings indicate that PhoB protein recognizes and binds to the pho box and activates transcription of the genes in the phosphate regulon.
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PMID:Regulation of the phosphate regulon of Escherichia coli. Activation of pstS transcription by PhoB protein in vitro. 305 25

The control region of the carAB operon, encoding carbamoylphosphate synthetase, comprises two tandem promoters (P1, upstream and P2, downstream) located 67 base-pairs apart and repressed respectively by pyrimidines and arginine. RNA polymerase and pure arginine repressor bind to the P2 region in mutually exclusive ways. Repressor protects the two adjacent palindromic ARG boxes overlapping P2 against DNase I. Binding of RNA polymerase to P1 is abnormal; the region protected against DNase I is shifted upstream by about 20 nucleotides with respect to the position expected from the transcription startpoint. This pattern is not due to interference with polymerase binding at P2, since it is observed also in the presence of repressor and on an isolated P1 region. Binding of RNA polymerase is relatively weak and heparin-sensitive suggesting that, in vivo, an ancillary factor is required to promote the formation of an open complex. S1 nuclease mapping experiments show that the simultaneous presence of pyrimidines and arginine represses the downstream arginine-specific promoter (P2) more efficiently than arginine alone. This effect is not due to a direct regulatory interaction between pyrimidines and P2, since it is not observed when P1 is inactivated by insertion mutations or partial deletion. It has been shown that transcription initiated at P1 can proceed even when arginine represses P2. We therefore suggest that P2 operator-arginine repressor complex is destabilized by RNA polymerase binding at P1 or transcription from P1. We describe a novel technique to select for expression-down mutants in a lac fusion context.
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PMID:Molecular interactions in the control region of the carAB operon encoding Escherichia coli carbamoylphosphate synthetase. 306 19

The gene for human liver-type arginase (EC 3.5.3.1), a urea cycle enzyme, was cloned and the structure was determined. This gene is 11.5 kilobases long and is split into 8 exons. The cap site was determined by nuclease S1 mapping and primer extension. A "TATA box"-like sequence is located 28 bases upstream from the cap site, and a sequence similar to the binding sites of the transcription factor CTF/NF1, a "CAAT box"-binding protein, is located 72 bases upstream. In the 5' end region, sequences resembling the glucocorticoid responsive elements, the cAMP responsive elements, and the enhancer core sequences are present. The immediately 5' flanking region of the human gene up to position -105 is 84% identical with the corresponding segment of the rat gene. In this region of the human gene, one DNase I-protected area and several hypersensitive cleavage sites were detected by footprint analysis, using nuclear extracts from the rat liver. The protected area contains the sequence similar to the binding sites of CTF/NF1 and also overlaps with the sequence resembling the glucocorticoid responsive elements.
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PMID:Human liver-type arginase gene: structure of the gene and analysis of the promoter region. 317 33

Mouse c-myb gene transcripts in various cells of haemopoietic origin were analysed using S1 nuclease and RNase mapping techniques and by Northern blotting. It was found that the prevalent 3.8-kb c-myb mRNA present in thymocytes, T cell leukaemias, myelomonocytic leukaemias, erythroleukaemias and myeloid stem cells was initiated at several cap sites mapping within a region 97-244 bp upstream from the protein coding sequence. Utilization of additional cap sites mapping further upstream was also observed in certain cells, most notably thymocytes, and this gave rise to RNA species (4.3-5.6 kb) larger than the presumptive mRNA. In contrast, myeloma cell c-myb transcripts, which are much less abundant than those in more immature haemopoietic cells, were found to be initiated at a restricted set of cap sites mapping 244-277 bp upstream of the coding sequence. Hence, these data suggest that the abundance of the c-myb mRNA may be regulated by a process involving selective utilization of mRNA cap sites. Sites hypersensitive to DNase I were associated with mRNA cap sites in cells that expressed c-myb.
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PMID:Multiple c-myb transcript cap sites are variously utilized in cells of mouse haemopoietic origin. 360 90

Splenic erythroblasts of mice infected with the anemia-inducing strain of Friend virus can be isolated in large numbers with less than 5% contamination with other cell types. In short-term culture, the isolated cells will initiate globin synthesis and undergo other aspects of terminal differentiation only if erythropoietin (EP) is added to the medium. An early effect of the hormone on these cells is stimulation of total RNA synthesis. EP also causes initiation of transcription of the beta-globin genes after a lag period of 4 to 6 h. By 6 h, the transcription rate of beta-globin RNA is enhanced threefold, and by 12 h, it is nearly maximal at ca. 20 times the level of control cells which received no EP. Transcription rates of alpha and beta-globin genes are approximately equal to each other throughout the period of terminal differentiation. In the splenic erythroblasts, the chromatin structure in the vicinity of the beta-major globin gene was analyzed with two nucleases during these transcription rate changes. No S1 nuclease-hypersensitive site is detectable near the gene. The beta-major gene is quite sensitive to DNase I in comparison with the albumin gene; however, the level of sensitivity is the same before EP addition as it is during maximal gene transcription after EP addition. Also, a hypersensitive site near the 5' cap site of the beta-major gene is quantitatively equivalent both before and after EP addition. Analysis of cytosine methylation at two sites upstream from the gene showed no changes upon induction of beta-globin gene transcription by EP. Thus, the initiation of beta-globin transcription by EP appears to be at some step after chromatin structural alteration such as synthesis, release, or activation of a specific transcription initiation factor.
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PMID:Control of globin gene transcription by erythropoietin in erythroblasts from friend virus-infected mice. 399 Jun 88

DNA fragments of the Streptomyces lividans plasmid pIJ101 have been tested for their ability to bind Streptomyces coelicolor RNA polymerase in vitro or to promote transcription in Streptomyces in vivo. One DNA fragment which does both was shown to encode a transcript which was expressed at low cell-density in cultures of pIJ101-containing cells. The transcript start was located on the DNA sequence of the fragment by nucleotide-primed RNA polymerase binding experiments and by S1 nuclease mapping. The pattern of DNase I protection, the sites of enhanced DNase I cleavage and the DNA sequence of the fragment suggest that the RNA polymerase holoenzyme form, which recognizes this promoter, is similar in its interaction with DNA to the major RNA polymerase of Escherichia coli. Regions showing 3/6 nucleotide homology with each of the -35 and -10 regions of the consensus sequence of E. coli promoters are present in the same positions relative to the transcript start. Symmetrical sequences which may be involved in the regulation of expression of the promoter and a potential polypeptide coding sequence can be identified.
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PMID:RNA polymerase-DNA interactions in Streptomyces. In vitro studies of a S. lividans plasmid promoter with S. coelicolor RNA polymerase. 404 37


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