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)

Injection of high concentrations of benzo[a]pyrene (10-125 micrograms BaP/g body weight) into young frogs (Xenopus laevis), fish (Gambusia affinis) or tissue culture cells (L 5178 Y) in a state of logarithmic growth causes alterations in DNA. In frogs these alterations reach a maximum at 60-90 min after application, then they decrease and become undetectable after 3 h. Within 8-12 days after the single BaP dose, a new wave of DNA alterations can again be detected. Parallel measurements of mixed function oxygenases (MFO) showed no short term activity changes within the first day. There was, however, an increase of activity starting on the 8th day following injection in frogs, and lasting until day 14. DNA alterations have been assessed by two methods. One is a modification of the alkaline filter elution method of Kohn et al. (1976) and is believed to determine the number of events leading to single strand breaks at alkaline pH. The other method determines the number of S1 nuclease sensitive sites in highly purified native DNA. Both results were found to be highly correlated. The effect does not seem to be caused by an impurity, unless an unknown, very minor, and extraordinarily active component is present. The effect in frogs is linked to nutritional status. Frogs fed a carbohydrate-rich, protein-poor diet do not give the fast response. The fast effect cannot be blocked by application of Actinomycin, Cordycepin, and Chloramphenicol injected either 30 min before or simultaneously with the BaP. Benzo(e)pyrene, a non-tumorigenic isomer of BaP, does not cause any effects at similar doses.
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PMID:Fast ephemeral DNA damage upon BaP injection. 632 Mar 64

We have analyzed a cloned beta O-thalassemia (beta O-thal) gene from a patient doubly heterozygous for hemoglobin Lepore and beta O-thalassemia. Studies of 3H-uridine incorporation into beta-globin mRNA in this patient's erythroblasts suggested an intranuclear defect in both beta and Lepore (delta beta) mRNA synthesis, as did S1 nuclease analysis of nuclear RNA. However, the nucleotide sequence of the beta O-thal gene revealed only a single base change in codon 39 (CAG----UAG), which created a premature translation termination codon. The 5' flanking sequence, including transcription promotor boxes and the mRNA initiation (CAP) site, were normal. The unexpected effect of this mutation on intranuclear beta-mRNA synthesis in vivo was studied by insertion of the cloned gene into a plasmid expression vector and transfection into tissue culture (COS-1) cells. beta-Globin mRNA produced by the transfected cells was assessed by S1 nuclease analysis. The beta O-39 thalassemia gene generated five- to tenfold less beta-mRNA than a normal beta-gene in both nuclear and cytoplasmic RNA, simulating the results observed in vivo. Moreover, the small amount of beta O-39 mRNA produced was as stable as normal beta-mRNA during an actinomycin D chase, ruling out rapid cytoplasmic turnover as a cause of the reduced accumulation. Cotransfection of the beta O-39 thalassemia gene with a mutant tyrosine suppressor tRNA gene resulted in restoration of the beta O-39 mRNA accumulation to near-normal levels. On the basis of these results, we suggest that the low levels of beta-mRNA known to exist in the common form of beta O-thalassemia, beta O-39 thalassemia, result from a lesion in transcription, or early posttranscriptional processes; the defect appears to be corrected by restoration of proper translational potential to the mutant mRNA, at least in a gene transfer-expression system in tissue-culture cells.
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PMID:Intranuclear defect in beta-globin mRNA accumulation due to a premature translation termination codon. 673 65

In Pseudomonas putida carrying the CAM plasmid, the operon (camDCAB) encoding enzymes involved in the degradation pathway of D-camphor is negatively regulated by the CamR protein, and camR is autorepressed. S1 nuclease mapping revealed that camDCAB and camR were divergently transcribed from overlapping promoters, the transcription start sites were separated by 11 bp, and transcriptions of the cam operon (camDCAB) and camR increased about 10- and 4-fold, respectively, immediately after addition of camphor. The transcriptions of camDCAB and camR were negatively regulated through the interaction of the CamR protein with the one operator located in the overlapping promoter region. In vitro transcription experiments were performed to characterize the regulation of cam genes. The camR promoter was initiated by P. putida RNA polymerase containing sigma 70, but transcription from the camDCAB promoter by sigma 70 holoenzyme was not observed. The purified CamR protein repressed in vitro transcription from the camR promoter. This repression was suppressed by camphor. The RNA polymerase binding region of the camR promoter was identified by using DNase I footprinting. In addition, footprinting studies revealed that the CamR protein and RNA polymerase coexisted on the promoter region in a joint nonproductive complex.
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PMID:Transcription of the cam operon and camR genes in Pseudomonas putida PpG1. 769 53

Overexpression of the Fli-1 gene has been shown to be involved in retrovirus-induced mouse tumors. Cloning of the 5' flanking sequence of the mouse and human Fli-1 exon 1 was performed. At least two major transcription initiation sites were localized respectively at 143 and 114 nucleotides upstream of the previously defined mouse Fli-1 cDNA 5' end. The sequences flanking the CAP sites show good conservation between human and mouse (94%). The promoter region contains a potential TATA box lying 30 bp from one of the major identified CAP sites. Several conserved elements, such as GATA, EBS, GC rich, AP-2, AP-3 elements and a repetition of GA were observed next to the two major CAP sites. Furthermore, this latter was shown to form a H-DNA structure in vitro by S1 nuclease sensitivity experiments. The highly conserved 5' non-translated region of exon 1 is predicted to form a very stable hairpin structure which could regulate the Fli-1 expression at the post-transcriptional level. In Cas-Br-E-induced tumors, all the proviruses are found clustered within 35 nucleotides directly upstream the Fli-1 ATG start codon, thus deleting the hairpin structure from the transcript. Promoter activity was tested using the CAT reporter gene transfected in mouse and human erythroid cell lines. No promoter activity could be detected with various mouse Fli-1 promoter-CAT constructs containing 600 bp of the 5' flanking region, the complete exon 1, the 5' end of intron 1 and/or retroviral LTR sequence. Constructions of the human homologue containing nearly 1.5 kbp of Fli-1 5' flanking region was also inactive in transfected cells. These results suggest that multiple levels of regulation might control the Fli-1 expression.
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PMID:Characterization of the human and mouse Fli-1 promoter regions. 867 8

The Pseudomonas aeruginosa homolog of the Escherichia coli global transcriptional regulator CRP (or CAP) was recently identified and designated Vfr (S. E. H. West, A. K. Sample, and L. J. Runyen-Janecky, J. Bacteriol. 176:7532-7542, 1994). Nucleotide sequence analysis of the region 5' to vfr identified a 423-bp open reading frame (ORF), which was designated orfX. The deduced amino acid sequence of ORFX was 53% identical and 87% similar to a divergent ORF of unknown function located 5' to the E. coli crp gene. When orfX was expressed from a phage T7 promoter in E. coli, a protein with an apparent molecular mass of approximately 18 kDa was produced. We constructed a chromosomal deletion of the region containing the 5' end of orfX (orfX'), vfr, and the 3' end of trpC (trpC') in P. aeruginosa strains PAO1 and PA103. The cloned vfr gene restored Vfr-dependent production of exotoxin A and protease in the PA103 orfX'-vfr-trpC' deletion mutant, suggesting that ORFX is not required for Vfr production or activity. To determine whether transcription of orfX and vfr are controlled by the same mechanisms that control transcription of the region of the divergent ORF (dorf) and of crp, we compared the vfr-orfX and crp-dorf intergenic regions. Using S1 nuclease analysis, we determined that the distance between the orfX and vfr transcriptional start sites was 105 bp. Thus, the P. aeruginosa orfX and vfr promoters are arranged in a back-to-back orientation rather than the face-to-face orientation of the dorf and crp promoters. A CRP recognition site is associated with each promoter in the crp-dorf intergenic region; binding of the CRP-cyclic AMP complex to the stronger dorf CRP recognition site activates transcription from the dorf promoter and represses transcription from the crp promoter. The vfr-orfX intergenic region does not contain an obvious CRP recognition site. In addition, vfr was not required for transcription of orfX. Unlike the dorf and crp mRNAs, the 5' ends of the orfX and vfr mRNAs were not complementary. Thus, the orfX mRNA cannot hybridize to the 5' end of the vfr mRNA to inhibit vfr transcription, a mechanism that has been postulated to control crp transcription in E. coli.
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PMID:A divergently transcribed open reading frame is located upstream of the Pseudomonas aeruginosa vfr gene, a homolog of Escherichia coli crp. 913 92

Investigation of the expression of the riboflavin (rib) genes, which are found immediately downstream of luxG in the lux operon in Photobacterium phosphoreum, provides more information relevant to the evolution of bioluminescence, as well as to the regulation of supply of flavin substrate for bacterial bioluminescence reactions. In order to answer the question of whether or not the transcriptions of lux and rib genes are integrated, a transcriptional termination assay was performed with P. phosphoreum DNA, containing the possible stem-loop structures, located in the intergenic region of luxF and luxE (OmegaA), of luxG and ribE (OmegaB), and downstream of ribA (OmegaC). The expression of the CAT (Chloramphenicol Acetyl Transferase) reporter gene was remarkably decreased upon the insertion of the stem-loop structure (OmegaC) into the strong lux promoter and the reporter gene. However, the insertion of the structure (OmegaB) into the intergenic region of the lux and the rib genes caused no significant change in expression from the CAT gene. In addition, the single stranded DNA in the same region was protected by the P. phosphoreum mRNA from the S1 nuclease protection assay. These results suggest that lux genes and rib genes are part of the same operon in P. phosphoreum.
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PMID:Coregulation of lux genes and riboflavin genes in bioluminescent bacteria of Photobacterium phosphoreum. 1545 47


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