Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.30.2 (endonuclease)
 18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

O6-Alkylguanine-DNA alkyltransferase (ATase) activity was determined in crude sonicates of tissues obtained from the F2 offspring of human ATase transgenic founder mice. In certain cases, samples were analyzed both before and after administration of zinc sulfate in the drinking water for 2 wk to upregulate the mouse metallothionein-1 promoter that controls the expression of the transgene. In liver samples obtained by partial hepatectomy, the ATase activities of nontransgenic mice ranged from 63 to 139 fmol/mg total protein (mean of 10 mice, 95.3 +/- 23 fmol/mg), whereas in positive transgenic mice, the range was from 503 to 2119 fmol/mg (mean of 10 mice, 963 +/- 475 fmol/mg). The difference between the mean ATase values for these two groups of mice is highly significant (P less than 0.001). All positive mice expressed ATase and in those examined using the human ATase coding sequence as a probe, isoschizomeric-restriction endonuclease digestion showed no evidence of cytosine methylation in the transgene. After zinc sulfate induction, the ATase levels in residual liver tissue were for the controls 84-191 fmol/mg (mean of 10 mice, 123 +/- 31.5 fmol/mg) and for positive mice 908-3273 fmol/mg (mean of 10 mice, 1960 +/- 724 fmol/mg). Induction thus caused a 1.4- to 3.2-fold increase in ATase activity in the tissues of individual transgenic mice (mean, 1.8-fold; P less than 0.003), with the greatest increase generally occurring in those mice that had the lowest preinduction levels. Hepatic ATase levels were thus increased up to 28 times higher in transgenic mice than in nontransgenic mice. When data from other groups of transgenic and nontransgenic mice (eight of each) was included and analyzed in an independent rather than paired fashion, the mean values for zinc-treated controls and transgenic mice, respectively, were 106 fmol/mg and 1415 fmol/mg, still a highly significant (P less than 0.001) difference. In two mice given a single intraperitoneal dose of cadmium chloride, hepatic ATase increased 2.1- and 4.9-fold, respectively. The effect of partial hepatectomy alone was also considered: for transgenic mice the mean ATase level increased from 453 to 661 fmol/mg protein after 48 h. In other offspring subjected to either unilateral nephrectomy or orchidectomy, induction of ATase activity by zinc sulfate was also seen in kidney (5.7- and 8.4-fold) and testis (1.7- and 3.1-fold), although these observations were made with small numbers of mice.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Tissue-specific expression and induction of human O6-alkylguanine-DNA alkyltransferase in transgenic mice. 150 42

Specific peptides of varying lengths were inserted between the two metal cluster domains of metallothionein (MT), which normally are spanned by only three amino acids, Lys-Lys-Ser. These interdomain expansions were made to test if such structural alterations would affect MT function. These constructs were engineered by inserting defined oligonucleotides of up to four tandem repeats of dodecanucleotides and hexanucleotides into an Alu-1 endonuclease cleavage site, which separates the two exonic regions of an MT-coding sequence from Chinese hamster ovary cells, MT-2. The native and altered sequences were cloned into a high expression Escherichia coli-yeast shuttle vector and used to transform yeast cells whose endogenous MT genes had been previously deleted. Using metal resistance as a biological marker, all constructs were shown to be functional in rendering the host cells resistant to either copper or cadmium. As the inserts, by nature of their amino acid sequence, could add flexibility to the otherwise compact molecule, the two domains apparently are active independently. The level of activity, however, diminished with the length of the insert. Determinations for copy number of the chimeric plasmids and MT mRNAs in the transformed cells showed that the replicational and transcriptional capacity of the long and short constructs were equivalent.
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PMID:Metallothioneins with interdomain hinges expanded by insertion mutagenesis. 218 35

Restriction endonuclease fragment length variations (RFLVs) were found through the use of cDNA probes for metallothionein genes 1 (Mt-1) and 2 (Mt-2) in the mouse. RFLVs were detected in restriction patterns generated by BglII and XbaI in the Mt-1 gene and by PvuII in the Mt-2 gene. All laboratory strains carry the Mt-1a and Mt-2a alleles. Among strains of wild origin, some Western European subspecies (Mus mus domesticus and M. m. brevirostris) also carry the Mt-1a and Mt-2a alleles. In contrast, a European subspecies (M. m. musculus) and the great majority of subspecies from East Asian countries (M. m. molossinus, Chinese mice of wild origin, and M. m. yamashinai) carry the Mt-1b and Mt-2b alleles. A domesticus strain from Bulgaria and two castaneus strains from Thailand and Philippines carry the intermediate combination of Mt-1b and Mt-2a alleles. Using the RFLVs, we mapped the Mt-1 and Mt-2 genes on chromosome 8, and they appear to be very closely linked since no recombination was observed between them in any of the mice examined. Data from three-point cross tests showed that the recombination frequencies are 4.31% between Os and Mt, 15.52% between Mt and Prt-2, and 19.83% between Os and Prt-2. The gene order of Os-Mt-1,Mt-2-Prt-2 has been confirmed.
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PMID:Restriction fragment length variations and chromosome mapping of two mouse metallothionein genes, Mt-1 and Mt-2. 257 52

Using a microinjection method (Rokkones et al. 1985) deoxyribonucleic acid was introduced into fertilized salmonid eggs. The survival rate after a 28 day period was 91% for injected eggs in comparison to non-injected controls. A gene construct containing the mouse metallothionein promoter fused to the human growth hormone structural gene was microinjected either as a supercoiled plasmid or as a linear sequence. In Southern blot analysis of both 5 and 73 day old dissected rainbow trout embryos, as well as in 1 year old Atlantic salmon, the mouse metallothionein human growth hormone gene sequence was detected together with the chromosomal DNA when micro-injected as plasmid or as linear DNA. After digestion with Bam HI restriction endonuclease, the human growth hormone gene was excised from the high molecular weight DNA fraction. Transcription into human growth hormone specific RNA, as well as translation and release of human growth hormone immunoreactive protein, could be demonstrated in early embryonic stages.
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PMID:Microinjection and expression of a mouse metallothionein human growth hormone fusion gene in fertilized salmonid eggs. 271 58

To study the factors essential for a functional restriction system, the PaeR7 restriction-modification system has been introduced and expressed in murine cells. Transfer of this system was accomplished in two steps. First, cells containing sufficient PaeR7 methylase to completely methylate the mouse genome were constructed. In the second step, the mouse metallothionein promoter-regulated, endonuclease expression vector linked to the hygromycin B resistance selection marker was used to transfect the high methylase-expressing cells. Sixty percent of the clones isolated contained PaeR7 endonuclease enzymatic activity. Transfected cells expressing both methylase and endonuclease were incapable of blocking infection by DNA viruses, and possible explanations are discussed.
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PMID:Introduction and expression of the bacterial PaeR7 restriction endonuclease gene in mouse cells containing the PaeR7 methylase. 285 May 39

The cytogenetic endpoints sister chromatid exchange (SCE) and chromosome aberrations are widely used as indicators of DNA damage induced by mutagenic carcinogens. Chromosome aberrations appear to result directly from DNA double-strand breaks, but the lesion(s) giving rise to SCE formation remains unknown. Most compounds that induce SCEs induce a spectrum of lesions in DNA. To investigate the role of double-strand breakage in SCE formation, we constructed a plasmid that gives rise to one specific lesion, a staggered-end ("cohesive") DNA double-strand break. This plasmid, designated pMENs, contains a selectable marker, neo, which is a bacterial gene for neomycin resistance, and the coding sequence for the bacterial restriction endonuclease EcoRI attached to the mouse metallothionein gene promoter. EcoRI recognizes G decreases AATTC sequences in DNA and makes DNA double-strand breaks with four nucleotides overhanging as staggered ends. Cells transfected with pMENS were resistant to the antibiotic G418 and contained an integrated copy of the EcoRI gene, detectable by DNA filter hybridization. The addition of the heavy metal CdSO4 resulted in the intracellular production of EcoRI, as measured by an anti-EcoRI antibody. Cytogenetic analysis after the addition of CdSO4 indicated a dramatic increase in the frequency of chromosome aberrations but very little effect on SCE frequency. Although there was some intercellular heterogeneity, these results confirm that DNA double-strand breaks do result in chromosome aberrations but that these breaks are not sufficient to give rise to SCE formation.
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PMID:Inducible expression and cytogenetic effects of the EcoRI restriction endonuclease in Chinese hamster ovary cells. 305 12

To address the role of transient torsional stress in transcription, we have utilized the regulated expression of HO endonuclease in yeast to create double-strand breaks in DNA templates in vivo at preselected sites. Linearization of circular minichromosomes, either 2 kb upstream or immediately downstream of a lacZ reporter gene controlled by the yeast metallothionein gene (CUP1) promoter, did not alter the copper induction profile of lacZ RNA transcripts compared to that of nonlinearized controls. Constructs site-specifically integrated into yeast chromosome II gave similar results. In vivo cross-linking with psoralen as a probe for negative DNA supercoiling demonstrated that template linearization efficiently dissipated DNA supercoiling induced by transcription. Therefore, the efficient transcription of linearized, relaxed templates found here demonstrates that transient torsional tension is not required for transcription of chromatin templates in yeast.
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PMID:Template topology and transcription: chromatin templates relaxed by localized linearization are transcriptionally active in yeast. 911 54

Cell death resulting from cadmium (Cd) intoxication has been confirmed to occur through apoptosis by morphological and biochemical studies. However it is still not clear whether Cd itself or metallothionein (MT) induced by Cd is the major factor responsible for the apoptosis. Although apoptosis is inducible by exposure of cells to various stimuli, the common pathway involved is generally accepted to be activation of endonucleases that induce internucleosomal cleavage of DNA, resulting in the 'ladder' formation observed upon agarose gel electrophoresis and the chromatin condensation seen by electron microscopy. Cd does not seem to activate the endonuclease in vitro. However, Cd itself can be associated with apoptosis through indirect oxidative stress by inhibition of antioxidant enzymes and possible interaction with zinc finger protein. In addition to the direct effect of Cd, MT appears to play dual roles in apoptosis induction: one as a Cd carrier by which Cd accumulates in the nucleus, and the other as an inhibitor of zinc finger proteins, which include transcriptional factors related to apoptosis such as the product of the apoptosis resistance gene A20. In this review, we demonstrated that the mode of cell death following Cd exposure is associated with intracellular movement of Cd and MT. A possible mechanism for Cd-induced apoptosis is also discussed.
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PMID:Apoptosis induced by cadmium. 1464 32

A complex network of RNA transcripts is generated from eukaryotic genomes, many of which are processed in unexpected ways. Here, we highlight how premature transcription termination events at protein-coding gene loci can simultaneously lead to the generation of short RNAs and attenuate production of full-length mRNA transcripts. We recently showed that the Integrator (Int) complex can be selectively recruited to protein-coding gene loci, including Drosophila metallothionein A (MtnA), where the IntS11 RNA endonuclease cleaves nascent transcripts near their 5' ends. Such premature termination events catalyzed by Integrator can repress the expression of some full-length mRNAs by more than 100-fold. Transcription at small nuclear RNA (snRNA) loci is likewise terminated by Integrator cleavage, but protein-coding and snRNA gene loci have notably distinct dependencies on Integrator subunits. Additional mechanisms that attenuate eukaryotic gene outputs via premature termination have been discovered, including by the cleavage and polyadenylation machinery in a manner controlled by U1 snRNP. These mechanisms appear to function broadly across the transcriptome. This suggests that synthesis of full-length transcripts is not always the default option and that premature termination events can lead to a variety of transcripts, some of which may have important and unexpected biological functions.
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PMID:Attenuation of Eukaryotic Protein-Coding Gene Expression via Premature Transcription Termination. 3210 32