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)

The upstream region of the rat CYP17 gene shows significant homology to the upstream regions of the bovine and human genes, 53 and 60 percent, respectively. The start site of transcription was determined by primer extension and S1 nuclease protection to be 41 base pairs (bp) upstream of the initiating methionine codon. Expression vectors were constructed by ligation of upstream sequences into promoterless chloramphenicol acetyl transferase (CAT) vectors. Transient transfection studies using primary cultures of rat Leydig cells indicate a strong cAMP-responsive element located within the -26/+65 region. Stimulation by cyclic AMP was abolished when sequences upstream of -264 were included in expression vectors. No significant expression was seen in Leydig cells in the absence of dbcAMP nor was there any expression in the presence or absence of dbcAMP in rat skin fibroblasts or in mouse adrenal (Y-1) cells in which CYP17 is not normally expressed. Three possible regulatory elements were found in the 5' upstream region: a CRE/ATF consensus sequence (GACGTCA) starting at position -57; a GRE consensus sequence (TGTTCT) starting at position -501; and a consensus sequence for AP-1 binding (TTAGTCA) starting at position -659. It was concluded that the CRE/ATF at -57 is not responsible for increased transcription in the presence of cyclic AMP.
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PMID:Cyclic AMP regulates expression of the rat gene for steroid 17 alpha-hydroxylase/C17-20 lyase P-450 (CYP17) in rat Leydig cells. 132 85

The src family gene lyn is expressed preferentially in B lymphocytes but very little in normal T lymphocytes. Transcription of the lyn gene in T lymphocytes was shown to be induced by the p40tax protein encoded by human T-cell lymphotropic virus type I. For determination of the mechanism of p40tax-mediated trans activation, the transcriptional promoter region of the lyn gene was characterized. By endonuclease S1 mapping, the transcriptional initiation sites were identified within the 770-bp EcoRI-SacI fragment of the 5'-terminal portion of the human lyn gene. This fragment showed promoter activity when placed upstream of the bacterial chloramphenicol acetyltransferase gene and transfected into various cell lines. Nucleotide sequence analysis revealed that the lyn promoter region contained four GC box-like sequences but not a TATA or CCAAT box. In addition, it contained sequences characteristic of a cyclic AMP-responsive element, octamer-binding motif, PEA3-like motifs, and NF kappa B-binding motif-like sequence. Mutational analysis suggested that the octamer-binding motif sequence is of primary importance for the lyn promoter activity but that the other elements are not. Cotransfection of various chloramphenicol acetyltransferase constructs containing different length of the lyn promoter together with p40tax expression plasmids into Jurkat T cells showed that the sequence responsible for p40tax-induced transcription is present around the transcription initiation sites.
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PMID:Characterization of the promoter region of the src family gene lyn and its trans activation by human T-cell leukemia virus type I-encoded p40tax. 150 84

The Klebsiella aerogenes gene maoA, which is involved in the synthesis of monoamine oxidase, was induced by tyramine and the related compounds, subjected to catabolite and ammonium ion repression, and cloned. The nucleotide sequence of the region involved in monoamine oxidase synthesis was determined. Two open reading frames, the maoA gene and a hitherto unknown gene (maoC), were found. These are located between a potential promoter sequence and a transcriptional terminator sequence. A region of the Escherichia coli chromosome that was highly homologous to the Klebsiella maoA gene was found. The potential maoA gene is located at 30.9 min on the E. coli chromosome. Analysis of the amino acid sequences of the first 11 amino acids from the N terminus of the purified monoamine oxidase agrees with those deduced from the nucleotide sequence of the maoA gene. The leader peptide extends over 30 amino acids and has the characteristics of a signal sequence. Primer extension and S1 nuclease mapping of transcripts generated in vivo suggests that the tyramine-induced mRNA starts at a site 62 bases upstream from the ATG initiation codon of the maoC gene. In the putative promoter region, a high degree of similarity to the consensus sequence for the binding site of cyclic AMP receptor protein was found. Thus, the mao region is composed of two cistrons, and the mao operon is regulated by monoamine compounds, glucose, and ammonium ions.
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PMID:A monoamine-regulated Klebsiella aerogenes operon containing the monoamine oxidase structural gene (maoA) and the maoC gene. 155 68

A simple procedure, involving heat-treatment, DEAE-Sephadex, AMP-Sepharose and Bio-Gel P-60 chromatography, was developed for the purification of S1 nuclease to homogeneity from commercially available Takadiastase powder. Chemical modification of the amino groups of purified S1 nuclease revealed that lysine is essential for single-stranded DNAase, RNAase and phosphomonoesterase activities associated with the enzyme. The kinetics of inactivation suggested the involvement of a single lysine residue in the active site of the enzyme. Additionally, lysine modification was accompanied by a concomitant loss of all the activities of the enzyme, indicating the presence of a common catalytic site responsible for the hydrolysis of single-stranded DNA, RNA and 3'-AMP. Substrate-protection and inhibitor-binding studies on enzyme modified with 2,4,6-trinitrobenzenesulphonic acid showed that lysine may be involved in the substrate binding.
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PMID:Active-site characterization of S1 nuclease. I. Affinity purification and influence of amino-group modification. 163 40

Two rat genomic clones, one for cytochrome P-450aldo and the other for P-450(11) beta, were isolated and characterized. The two genes, encoding structurally homologous proteins, were closely similar in their intron-exon organizations. Their 5'-flanking regions, however, contained only a few homologous regions. A putative cyclic AMP responsive element, TGACGTGA, was found in the P-450aldo gene, but this sequence was altered at two positions in the P-450(11) beta gene. S1 nuclease protection assay revealed a single transcription initiation site for the P-450aldo gene, while multiple sites were found for the P-450(11) beta gene. These results suggest that transcriptional regulation of the rat P-450aldo and P-450(11) beta genes is due to differences in the sequences of their 5'-flanking regions.
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PMID:Structural differences in 5'-flanking regions of rat cytochrome P-450aldo and P-450(11) beta genes. 195 71

The ugp operon of Escherichia coli includes genes involved in the uptake of sn-glycerol-3-phosphate and glycerophosphoryl diesters and belongs to the pho regulon which is induced by phosphate limitation. This operon has two transcriptional initiation sites, as determined by S1 nuclease mapping of the in vivo transcripts. The downstream promoter has multiple copies of the pho box, the consensus sequence shared by the pho promoters; the upstream promoter has a consensus sequence for the promoters regulated by cyclic AMP and its receptor protein, CRP. PhoB protein, which is the transcriptional activator for the pho regulon, protected the regulatory region with the pho boxes in DNase I footprinting experiments and activated transcription from the downstream promoter in vitro. Studies with transcriptional fusions between ugp and a promoterless gene for chloramphenicol acetyltransferase show that the upstream promoter is induced by carbon starvation in a manner that required the cya and crp genes. PhoB protein may act as a repressor for this upstream promoter, which also overlaps the upstream third pho box. The downstream promoter was induced by phosphate starvation and requires the PhoB protein for its activation as do the other pho regulon promoters. These results suggest that the two promoters function alternately in responding to phosphate or carbon starvation, thus providing the cell with a means to adapt to these physiological stresses.
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PMID:Dual regulation of the ugp operon by phosphate and carbon starvation at two interspaced promoters. 198 50

A catabolite-sensitive promoter was found to be involved in transcription of the heat shock regulatory gene rpoH encoding the sigma 32 protein. Expression of lacZ from the operon fusion, rpoHp-lacZ, was partially inhibited by glucose added to the broth medium. Dissection of the rpoH promoter region allowed us to localize the glucose-sensitive promoter to the 110-base-pair (bp) segment directly upstream of the rpoH coding region. Experiments on lacZ expression from the set of fusions in cya (adenylate cyclase) and crp (cyclic AMP [cAMP] receptor protein) mutants also supported the involvement of a catabolite-sensitive promoter. Analysis of rpoH mRNAs by S1 nuclease protection experiments led us to identify a novel promoter, designated P5, that is regulated by cAMP and the cAMP receptor protein. Studies of rpoH transcription in vitro demonstrated that RNA polymerase-sigma 70 can transcribe from the P5 promoter only in the presence of cAMP and its receptor protein. The 5' ends of P5 transcripts obtained in vivo and in vitro were found to be at 61 to 62 bp upstream of the initiation codon, and a putative binding sequence for the cAMP receptor protein was found at 38 to 39 bp further upstream. Transcription from the P5 promoter is increased by the addition of ethanol to the growth medium; however, the increase is greater in the presence of glucose than in its absence. These results add a new dimension to the transcriptional control of rpoH and to the regulation of the heat shock response in Escherichia coli.
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PMID:Transcriptional regulation of the heat shock regulatory gene rpoH in Escherichia coli: involvement of a novel catabolite-sensitive promoter. 213 50

Genomic clones containing the 5'-terminal portion of the human CRE-BP1 gene that encodes transcriptional regulator binding to the cyclic AMP response element (CRE) were isolated. Multiple transcriptional start sites in the promoter region were identified by nuclease S1 mapping and primer extension analysis. By DNase I footprinting with use of purified transcription factor Sp1 and nuclear extracts prepared from HeLa cells, 11 Sp1-binding sites, two CCAAT sequences, two CREs, and three unknown factor recognition elements were found. Transfection of chimeric chloramphenicol acetyltransferase plasmids containing various deletions of the promoter into CV-1 cells indicated that the region between nucleotides -50 and 90, which contained three Sp1-binding sites and one CRE, was sufficient for basal promoter activity. These results suggest that multiple sequence-specific DNA-binding proteins may control the expression of the CRE-BP1 gene, although Sp1 seems to be important for the basal promoter activity.
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PMID:Promoter region of the human CRE-BP1 gene encoding the transcriptional regulator binding to the cyclic AMP response element. 214 72

The identities of two cloned, arabinose-inducible promoters were tested by hybridizing promoter DNA fragments with restriction digests of chromosomal DNA containing Mudlac phage inserted in either araFGH or in araE transport operons. One promoter, thought to be araE, is within 10(3) base-pairs of a Mudlac insertion in the araE gene. The second promoter was not found within several thousand base-pairs of either of the known transport genes. This promoter is now named araPJ (araJ). The DNA sequence of the fragment containing the araFGH promoter was determined. The start site of transcription in vivo was located to within +/- 1 base-pair (bp) by S1 nuclease mapping. DNase 1 footprinting revealed that, in comparison with the araBAD and araE promoters, the locations of the AraC and cyclic AMP receptor protein (CRP) binding sites are reversed with CRP lying between AraC and RNA polymerase. The central location of the CRP binding site may explain why the araFGH promoter is more catabolite sensitive than the other ara promoters. AraC and CRP were both required for maximal transcription in vitro, although a low level of transcription was detected with CRP alone. S1 nuclease mapping of mRNA-DNA hybrids from the araJ promoter located the transcription start point to within #/- 3 bp, and demonstrates that the promoter is dependent upon AraC protein and CRP in vivo. DNase footprinting showed that the location of the AraC protein binding site on araJ is adjacent to the RNA polymerase site, as seen at the araBAD and araE promoters. Two CRP sites were observed; one is upstream from the AraC site and one is downstream from the transcription start site.
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PMID:Characterization of the Escherichia coli araFGH and araJ promoters. 223 17

The tsx gene of Escherichia coli encodes an outer membrane protein, Tsx, which constitutes the receptor for colicin K and bacteriophage T6, and functions as a substrate-specific channel for nucleosides and deoxynucleosides. The mini-Mu element pEG5005 was used to prepare a gene bank in vivo, and this bank was used to identify T6-sensitive strains carrying the cloned tsx gene. Subcloning of the tsx gene into the multicopy plasmid, pBR322, resulted in a strong overproduction of Tsx. The sequence of a 1477-bp DNA segment containing tsx and its flanking regions was determined. An open reading frame (ORF) was found which was followed by a pair of repetitive extragenic palindromic sequences. This ORF translated into a protein of 294 amino acids (aa), the first 22 aa of which showed the characteristic features of a bacterial signal sequence peptide. The putative mature form of Tsx is composed of 272 aa with a calculated Mr of 31418. The aa sequence of Tsx shows an even distribution of charged residues (52 aa) and lacks extensive hydrophobic stretches. No significant homologies of Tsx to the channel-forming proteins OmpC, OmpF, PhoE and LamB from the E. coli outer membrane were detected. Using nuclease S1, we identified two transcription start points for the tsx mRNA which were separated by approx. 150 bp. Genetic data suggest that the synthesis of the larger mRNA species is directed by a weak promoter (P1) that is controlled by the DeoR repressor, whereas the smaller mRNA species is directed by the main promoter P2, which is negatively controlled by the CytR repressor and positively affected by the cyclic AMP/catabolite activator protein complex.
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PMID:Analysis of the tsx gene, which encodes a nucleoside-specific channel-forming protein (Tsx) in the outer membrane of Escherichia coli. 226 60


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