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:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The influence of terminal differentiation on UV-induced DNA damage and its repair in transcriptionally active and inactive genomic sequences was investigated using the murine 3T3-T proadipocyte cell culture system. Actively cycling 3T3-T cells terminally differentiate into adipocytes after exposure to media containing platelet-depleted human plasma. Suitable DNA fragments were analyzed from four genes: beta-actin, adenosine deaminase, dihydrofolate reductase, and lipoprotein lipase. As a result of 3T3-T cell differentiation, lipoprotein lipase and beta-actin expression was modified, whereas adenosine deaminase and dihydrofolate reductase expression was not affected. A DNA fragment representing the transcriptionally inactive locus 70-38 was also evaluated. UV-induced cyclobutane pyrimidine dimers, detected as UV-specific endonuclease-sensitive sites, in each fragment increased linearly as a function of UV dose (0-20 J/m2) independently of gene expression or differentiation. Sequence-specific repair of dimers was measured in stem and terminally differentiated 3T3-T cells after UV irradiation (10 J/m2). For undifferentiated stem cells, the rate and extent of dimer repair was higher in the actively transcribed adenosine deaminase and dihydrofolate reductase genes than in the inactive lipoprotein lipase or 70-38 fragments, the greater difference being observed in the first 8 h post-UV irradiation. In contrast, similar dimer repair rates were found for each DNA fragment in terminally differentiated 3T3-T cells. These data suggest that cellular differentiation is accompanied by a loss of heterogeneity in intragenomic DNA repair.
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PMID:Loss of intragenomic DNA repair heterogeneity with cellular differentiation. 193 6

The promoter region of the IL2R alpha gene 5' flanking sequence contains enhancer elements crucial for binding nuclear factors which upregulate transcription following T lymphocyte activation. A 3' exonuclease resistant oligonucleotide (3'A-IL28p, terminated by a free amine group at its 3' end) was designed to bind to the IL2R alpha promoter region from -273 to -246, forming a collinear triplex spanning the kappa B enhancer (-266 to -256) as well as most of the serum response element (CArG box, -251 to -244). The binding site specificity of this oligonucleotide was demonstrated in electrophoretic mobility shift assays and by inhibition of restriction endonuclease (HinfI) cleavage within the segment of the target DNA predicted to form a triplex with the oligonucleotide. Intact normal lymphocytes, preincubated for 2h with 3'A-IL28p, accumulated less IL2Ralpha mRNA relative to other mRNAs (c-myc, beta-actin, IL2R beta, IL-6) for up to 12h after PHA stimulation, than did lymphocytes treated with a control oligomer of similar composition but different sequence. Nuclear run-on studies demonstrated that the rate of IL2R alpha mRNA synthesis relative to c-myc and beta-actin was also selectively diminished by treatment with 3'A-IL28p. These experiments suggest that transcription of individual genes can be selectively modulated in living cells by sequence specific collinear triplex formation in regulatory enhancer sequences.
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PMID:Oligonucleotide inhibition of IL2R alpha mRNA transcription by promoter region collinear triplex formation in lymphocytes. 206 58

A beta-actin gene of carp (Cyprinus carpio) was isolated from a genomic EMBL3 library. The nucleotide sequence of the gene indicates six exons spanning 3.6 kb. Southern blot hybridization of restriction endonuclease digests of carp genomic DNA indicate that there are two copies of the beta-actin isotype and several other species of actin genes. The transcriptional start site is 85 bp and 24 bp downstream respectively from consensus CCAAT and TATA promoter elements. The organization of the carp beta-actin gene is identical to that of chicken, human, and rat genes in terms of size, exon/intron locations and junctions and in having a translationally silent first exon. The fish gene is 90% and 99% conserved at the nucleotide and amino acid levels, respectively, with land vertebrate beta-actin genes. Northern blot analysis of beta-actin gene expression indicated that the gene is highly expressed in brain, less so in muscle, and much less so in liver cells. The putative beta-actin proximal promoter of carp, identified by the conservation of known actin regulatory sequences, is transcriptionally active in both mammalian and piscine cells.
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PMID:Isolation and characterization of beta-actin gene of carp (Cyprinus carpio). 213 83

The limited DNA-excision repair in UV-irradiated nondividing fibroblasts from xeroderma pigmentosum complementation group C (XP-C) occurs in localized chromatin regions generating large DNA segments (at least 30-70 kb) free of pyrimidine dimers. A genomic fraction enriched for this DNA was isolated on the basis of the larger size of the repaired fragments after UV-endonuclease treatment and screened for specific genes. It contains more copies per microgram DNA of two transcriptionally active genes, beta-actin and dihydrofolate reductase, compared to the remaining DNA but an equal number of copies per microgram DNA of an inactive locus termed 754. We confirmed that the active genes were preferentially repaired by measuring the removal of pyrimidine dimers from specific genomic restriction fragments comprising these sequences. These results mean that a unique set of relatively large chromatin domains are repaired in nondividing XP-C cells, even though most of the DNA remains unrepaired. The repaired domains may be those containing the active genes. This specific repair may account for the relatively high UV-resistance of the nondividing cells. In normal cells, a very rapid repair of a restriction fragment containing the beta-actin gene and slow repair of the 754-containing fragment was detected indicating that a similar domain-oriented repair process also exists in these cells. These results are consistent with the previously discovered rapid repair of active genes compared to bulk DNA. Separate damage-recognition systems may exist in human cells for chromatin domains that contain transcribed regions and those that contain no transcribed regions. The latter system may be deficient in XP-C.
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PMID:Selective repair of specific chromatin domains in UV-irradiated cells from xeroderma pigmentosum complementation group C. 234 4

High molecular weight DNA from pleroceroid larvae of the tapeworm Spirometra mansonoides was purified from isolated nuclei by conventional techniques. The DNA so isolated has a melting temperature (Tm) of 87 degrees C and a guanine plus cytosine (G/C) content of 44%. 5-Methyl cytosine could not be detected in plerocercoid DNA by HPLC analysis of DNA hydrolysates, by radiolabeling 5'-termini of MspI digests with polynucleotide kinase, or by comparing restriction patterns generated by MspI and HpaII. Renaturation kinetics demonstrated that the genome of S. mansonoides contains repetitive as well as single copy sequences and has a genome size estimated at approx. 1.6 X 10(9) bp. Hybridization was carried out between plerocercoid DNA and cDNAs for human beta-actin, alpha-tubulin and growth hormone (hGH). Rationale for this analysis was based on known homologies among actin and tubulin genes in numerous species and on apparent similarities between hGH and a plerocercoid growth factor that may be reflected in similar DNA sequence. Scanning densitometry of dot blots demonstrated that the hGH probe annealed to the same extent at low stringency (1 M NaCl, 55 degrees C) to DNA from plerocercoids, rat liver and chicken erythrocytes; but this interaction was less than to DNA from human lymphocytes, calf thymus and mouse skin. Similar results were obtained when restriction endonuclease digests of these DNAs were analyzed by Southern transfer. Little or no hybridization of the growth hormone probe to plerocercoid DNA was evident at higher stringency (1 M NaCl, 65 degrees C). In contrast, human tubulin and actin probes showed extensive hybridization to pleroceroid restriction fragments under the high stringency conditions.
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PMID:Molecular characterization of the Spirometra mansonoides genome: renaturation kinetics, methylation, and hybridization to human cDNA probes. 236 5

RNA synthesis in K-562 human erythroleukemia cells was markedly curtailed by exposure to the uridine analogue 5-fluorouridine (FUrd). The inhibition of ribosomal RNA synthesis was accompanied by rapid declines in the steady-state levels of several, but not all, mRNAs, including gamma-globin mRNA. In this report, we demonstrate that gamma-globin mRNA species were decreased by as much as 40% within 2 hr of exposure to micromolar concentrations of FUrd. The decline in gamma-globin mRNA occurred at a rate that outstripped the normal rate of degradation of this mRNA by a factor of 25. The decline in cytoplasmic mRNA was not mirrored in the nucleus; northern blotting revealed that pre-mRNA levels were not reduced. Nuclease protection analyses of precursors from FUrd treated and untreated control cells did not reveal any qualitative differences. Thus, the decrease was not accounted for by drug-induced inhibition of new gamma-globin mRNA synthesis or misincorporation but must have been due to an FUrd-induced increase in gamma-globin mRNA degradation. Drug-induced instability of RNA was not a generalized feature of FUrd exposure, since neither beta-actin mRNA nor cytoplasmic rRNA, whose stabilities in untreated cells are similar to that of gamma-globin mRNA, were affected. Furthermore, the instability of gamma-globin mRNA did not decrease globin protein levels, presumably because the stability of the protein was not altered. The mechanism by which specific increased degradation of gamma-globin mRNA occurred is unknown, but it may have been due to the activation of cytoplasmic endonuclease.
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PMID:Reductions in gamma-globin mRNA levels restricted to the cytoplasm of 5-fluorouridine treated K-562 human erythroleukemia cells. 236 50

We have constructed a mammalian expression vector consisting of 3 kilobases of the human beta-actin gene 5' flanking sequence plus 5' untranslated region and intervening sequence 1 linked at the 3' splice site to a short DNA polylinker sequence containing unique Sal I, HindIII, and BamHI restriction endonuclease sites followed by a simian virus 40 (SV40) polyadenylylation signal. Two derivatives, containing the selection markers obtained from pSV2gpt or pSV2neo, were also generated. We find that the promoter activity of this vector is a great or greater than that of the SV40 early promoter in a variety of human and rodent cells. The vector was used to generate gamma-actin and beta-tubulin antisense transcripts in human fibroblast cell lines. The antisense transcripts accumulate to levels comparable with that of the highly abundant gamma-actin and beta-tubulin mRNAs.
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PMID:A human beta-actin expression vector system directs high-level accumulation of antisense transcripts. 244 31

The synthesis and steady state level of immediate early vaccinia virus-specific RNAs in interferon-treated chick embryo fibroblasts were determined by blot hybridization analysis using the cloned restriction endonuclease fragment pEJ 18 containing the gene of vaccinia virus WR-specific DNA polymerase as a probe. Even though early vaccinia virus WR RNA was still synthesized, accumulation of immediate early viral RNAs was strongly inhibited. Accumulation of beta-actin RNA was not affected. This indicated an enhanced degradation of vaccinia virus WR-specific early RNAs in interferon-treated chick embryo fibroblasts. This notion was supported by Northern blot analysis which revealed degradation of residual RNA of vaccinia virus WR-specific DNA polymerase. In contrast to interferon-treated mouse L 929 cells, ribosomal RNA is not degraded in interferon-treated vaccinia WR-infected chick embryo fibroblasts.
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PMID:Increased turnover of vaccinia virus-specific immediate early RNAs in interferon-treated chick embryo fibroblasts. 244 63

Israeli Holstein-Friesian dairy bulls were screened for restriction fragment length polymorphisms by hybridizing cloned DNA probes for bovine growth hormone, for chymosin, and for rat muscle beta-actin to restriction endonuclease-digested DNA immobilized on nitrocellulose filters. The population proved to be polymorphic at the growth hormone locus, with evidence consistent with the phenotypes being inherited in allelic fashion. A low level of polymorphism was also observed at one of the beta-actin gene family loci. The chymosin locus was monomorphic with the restriction enzymes utilized. The results illustrate the power of restriction fragment length polymorphism methodology in visualizing genetic variability in dairy cattle populations.
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PMID:Restriction fragment length polymorphism among Israeli Holstein-Friesian dairy bulls. 301 50

Influenza virus infection has adverse effects on the metabolism of two representative RNA polymerase II transcripts in chicken embryo fibroblasts, those coding for beta-actin and for avian leukosis virus (ALV) proteins. Proviral ALV DNA was integrated into host cell DNA by prior infection with ALV. Within 1 h after influenza virus infection, the rate of transcription of beta-actin and ALV sequences decreased 40 to 60%, as determined by labeling the cells for 5 min with [3H]uridine and by in vitro, runoff assays with isolated nuclei. The transcripts that continued to be synthesized did not appear in the cytoplasm as mature mRNAs, and the kinetics of labeling of these transcripts strongly suggest that they were degraded in the nucleus. By S1 endonuclease assay, it was confirmed that nuclear ALV transcripts disappeared very early after infection, already decreasing ca. 80% by 1 h postinfection. A plausible explanation for this nuclear degradation is that the viral cap-dependent endonuclease in the nucleus cleaves the 5' ends of new polymerase II transcripts, rendering the resulting decapped RNAs susceptible to hydrolysis by cellular nucleases. In contrast to the nuclear transcripts, cytoplasmic beta-actin and ALV mRNAs, which are synthesized before infection, were more stable and did not decrease in amount until after 3 h postinfection. Similar stability of cytoplasmic host cell mRNAs was observed in infected HeLa cells, in which the levels of actin mRNA and two HeLa cell mRNAs (pHe 7 and pHe 28) remained at undiminished levels for 3 h of infection and decreased only slightly by 4.5 h postinfection. The cytoplasmic actin and pHe 7 mRNAs isolated from infected HeLa cells were shown to be translated in reticulocyte extracts in vitro, indicating that host mRNAs were not inactivated by a virus-induced modification. Despite the continued presence of high levels of functional host cell mRNAs, host cell protein synthesis was effectively shut off by about 3 h postinfection in both chicken embryo fibroblasts and HeLa cells. These results are consistent with the establishment of an influenza virus-specific translational system that selectively translates viral and not host mRNAs.
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PMID:Metabolism and expression of RNA polymerase II transcripts in influenza virus-infected cells. 609 46


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