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)

Differential accessibility to DNA in tumor cell chromatin is important to growth, differentiation apoptosis, and the targeting of DNA modifying drugs. We now show that endonuclease accessibility to DNA in the nuclei of A431 human carcinoma cells is increased within 90 min by nontoxic nanomolar levels of okadaic acid, known to inhibit protein phosphatase 2A. This genomic hypersensitivity was partly enhanced by joint treatment with epidermal growth factor and okadaic acid but did not appear without the latter. Nuclei with greater DNA susceptibility showed a decrease in M(r) 80,000 DNA binding protein doublet specific for dAT-rich sequences concurrent with the "apparent" hyperphosphorylation of a M(r) 70,000 nuclear matrix protein. We propose that some of the tumor-promoting effects of okadaic acid may be partly associated with its ability to promote genomic susceptibility.
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PMID:Accessibility to DNA in carcinoma chromatin is promoted by nanomolar okadaic acid: effect on AT-rich DNA binding proteins. 133 Feb 92

Using Northern blot technique, the oncogene expression in normal pancreatic tissue and human pancreatic carcinoma PC-2 and PC-3 cell lines was studied. Four oncogene probes (c-N-ras, c-ki-ras, c-myc and c-fos) consisting of recovered endonuclease digested fragments were nick translated. After hybridization and autoradiography, none of the four oncogenes was expressed in the normal human pancreatic tissue, but the human pancreatic carcinoma PC-2 and PC-3 cell lines expressed the c-myc and c-ki-ras genes. Their transcripts were 2.7 and 6.0 kb, respectively. Expression of the other two oncogenes (c-N-ras and c-fos) was not detected. The results of this study together with those reported in the literature verify the fact that the expression of c-myc and c-ki-ras oncogenes may play a very important role in the development of human pancreatic carcinoma.
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PMID:[Expression of c-myc and c-ki-ras oncogene in human pancreatic carcinoma]. 139 43

The open reading frames of the phosphoprotein pp58 (BMRFI) and the deoxyribonuclease (BGLF5) of the Epstein-Barr-virus (EBV) strain M-ABA were cloned in the baculovirus expression vectors pAc373 and pAc360 and expressed in the Spodoptera frugiperda (SF158) insect cells. The recombinant phosphoprotein pp58 expressed in SF158 cells was recognized by the anti-pp58 rabbit anti-sera which were generated by immunizing rabbits with a TrpE-BMRFI fusion protein expressed in E. coli. DNA-cellulose chromatography showed that the recombinant pp58 exhibited DNA-binding activities. Immunofluorescence, immunoblot and ELISA analysis indicated that sera from patients with nasopharyngeal carcinoma (NPC) contained antibodies against pp58. The recombinant EBV DNase expressed in SF158 cells was recognized by the anti-EBV DNase rabbit anti-sera which were generated by immunizing rabbits with a TrpE-C-terminal part of BGLF5 fusion protein expressed in E. coli. The anti-EBV DNase rabbit anti-sera recognized also a protein of about 52 kDa in the EBV-harboring human B-cell lines Raji, Jijoye, B95-8, M-ABA and BL74 induced by TPA and n-butyrate. The recombinant EBV DNase exhibited exonuclease and endonuclease activities, a requirement for magnesium, and a high pH optimum (8.0). Its enzyme activities could be inhibited by sera from NPC patients and anti-EBV DNase rabbit anti-sera. Comparable studies of Raji EBV-DNase and recombinant EBV-DNase implied that recombinant EBV-DNase could also be used in the enzyme activity assay for the detection of NPC. In contrast to the enzyme inhibition test, immunofluorescence and immunoblot analysis demonstrated that the recombinant EBV DNase exhibited only a weak immunological reaction with NPC sera.
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PMID:Immunological characterization of the Epstein-Barr virus phosphoprotein PP58 and deoxyribonuclease expressed in the baculovirus expression system. 165 Mar 30

Chromosome G-banding analysis of two human mammary carcinoma cell lines, Elco and MCF-7, showed the existence of two X chromosomes in both cell lines. To determine the state of activity of the X chromosomes, a methylation-sensitive restriction endonuclease, HpaII, was used to distinguish the active X from the hypermethylated, inactive X chromosome with a probe for the phosphogalactokinase locus by Southern blot hybridization. DNA digested with the restriction enzymes PstI and BstXI showed a band at either 1.05 or 0.9 kilobases. After HpaII digestion, a 50% reduction in intensity was observed in the female controls, whereas total reduction of the band was observed for the tumor cell lines and the male control. This indicates the absence of an inactive X and the presence of only active X chromosomes in the mammary carcinoma cell lines and the male control. To investigate the mechanisms involved in the alteration of the X chromosome composition and activity, restriction fragment length polymorphism analyses of seven additional X chromosome markers (L1.28, DX13, p52A, pX65H7, L782, pA13.RI, and pXG-12) were performed on the DNA isolated from the tumor cells and controls. Heterozygosity for at least one of the seven markers was detected in the six female controls whereas only homozygosity was detected for each marker in the tumor cell lines and the male control. These results indicate that the two active X chromosomes identified in each of the two tumor cell lines are identical, resulting from duplication or nondisjunction of the active X and loss of the inactive X chromosome.
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PMID:Two identical active X chromosomes in human mammary carcinoma cells. 197 Nov 94

The use of Mabs for the detection and treatment of human carcinoma lesions can still be regarded in its infancy. As with other new approaches to cancer therapy, several conceptual as well as real problems exist when designing clinical protocols for Mab-directed immunotherapy. From the Mab standpoint, studies using the intact IgG have shown that, in a majority of patients injected with IgG, human anti-mouse IgG antibodies develop that hamper the effectiveness of subsequent antibody administration. It is believed that the human anti-mouse antibody response is directed against the Fc region of the IgG molecule. The elimination of this region through fractionation of the Mab to obtain the minimum binding site could result in a less immunogenic molecule. Another approach aimed at reducing the immunogenicity of the Mab would be to clone the genes encoding for individual Mabs, reduce them via restriction endonuclease techniques, and insert human immunoglobulin constant regions. The resulting chimeric antibodies are believed to reduce the development of human anti-mouse antibodies. Effective Mab therapy of human tumor lesions may also be achieved through the recruitment of a portion of the host's immunologic defense system. An example is the use of anti-idiotype Mabs that use as immunogen a Mab to a tumor antigen. The anti-idiotype antibodies are selected for binding to the antigen binding, or idiotype, region of the first Mab. The binding sites of the new anti-idiotype Mabs should reflect the 'internal image' of the original antigen. The anti-idiotype antibodies may be used to immunize patients (i.e., vaccines) in an attempt to mount an active immune response against the antigen-positive tumor cells. Recent studies have shown a synergism between interferon-alpha and an anti-idiotype Mab for the in-vivo antitumor activity in a murine B-cell lymphoma experimental model. Whether an interferon-mediated increase in the tumor antigen or the Fc receptor was part of the synergism was not investigated. Mabs alone have also been shown to elicit cytotoxic activity in vitro and tumoricidal activity in vivo. Antibodies of the IgG2a isotype can direct macrophage-mediated cytotoxicity. These studies revealed the importance of the number of antibody sites per cell as well as the number of cells that bind the IgG2a Mab, thus suggesting a 'threshold' requirement for the demonstration of effective tumor cell lysis in vitro and in vivo.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Augmentation of tumor antigen expression by recombinant human interferons: enhanced targeting of monoclonal antibodies to carcinomas. 197 58

Tissues stored as paraffin blocks are a potential source of DNA for retrospective clinicogenetic analysis. To assess the feasibility of Southern blot analysis, DNA extracted from paraffin blocks was compared with DNA obtained from fresh-frozen controls of the same tissues. Sections 50-100 microns thick cut from paraffin blocks of 11 normal tissues, 18 lymphoid lesions, and 9 gastric carcinoma samples were deparaffinized and incubated at 45 degrees C for 48 to 72 h in a sodium dodecyl sulfate (SDS)/proteinase K solution. Following organic extraction, alcohol precipitation, restriction endonuclease digestion, and gel electrophoresis, DNA was transferred to nylon membranes. 32P-labelled DNA probes for the immunoglobulin heavy-chain locus and T-cell receptor beta-chain gene were hybridized to the normal tissue and lymphoid samples; the gastric cancers were probed for the HER-2/neu protooncogene. Intact DNA was obtained from the majority of formalin-fixed samples, yielding results qualitatively similar to those from fresh tissues. Degradation is the most significant problem in analyzing DNA extracted from paraffin blocks and compromises accurate quantitation. DNA analysis using paraffin-embedded tissue has potential clinical and research applications and may be a particularly useful way to study gene abnormalities in unusual tumors infrequently available as fresh specimens.
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PMID:Extraction of DNA from paraffin blocks for Southern blot analysis. 198 65

Allelic deletions of the p53 gene previously were demonstrated by Southern hybridization to occur in high frequency in sporadic colon carcinomas and in a variety of other human tumors. We have examined the frequency of allelic loss of the p53 gene in carcinoma and dysplasia arising in patients with chronic ulcerative colitis who are heterozygous for the codon 72 polymorphism in exon 4 of the p53 gene. Cells derived from carcinoma and dysplasia specimens from 10 patients who were heterozygous at this locus were sorted by flow cytometry on the basis of DNA content. The p53 exon 4 region was amplified from diploid and aneuploid populations, via a polymerase chain reaction (PCR), and digested with BstUI. Three of three carcinomas, four of six dysplasias, and one patient who was indefinite for dysplasia demonstrated evidence of allelic loss of the p53 gene. Seven of ten cases of sporadic colon carcinoma, analyzed for comparative purposes, exhibited loss of a p53 allele. These results demonstrate that PCR analysis, followed by restriction endonuclease digestion of a polymorphic locus, can provide a rapid, definitive method for analyzing loss of heterozygosity in small numbers of cells from colonic mucosa. Such loss precedes cancer in ulcerative colitis and can be present in its earliest histologically identifiable precursor.
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PMID:Frequent loss of a p53 allele in carcinomas and their precursors in ulcerative colitis. 204 25

A cDNA corresponding to the BGLF5 open reading frame of the Epstein-Barr virus (EBV) genome and coding for an early DNase was inserted into the procaryotic expression vector pKK223-3. One bacterial clone producing the expected 52-kilodalton DNase was used as a source of EBV DNase. The 52-kilodalton Dnase was purified in the active form to near homogeneity by ammonium sulfate precipitation and successive chromatographies on phosphocellulose, DNA-cellulose, and gel filtration columns. The purified enzyme exhibited both exonuclease and endonuclease activities, an absolute requirement for divalent cations, an alkaline pH preference, and a typical residual activity in presence of 300 mM KCl. Moreover, the enzyme was specifically inhibited by human sera with high antibody titers to EBV early antigens. These properties are similar to those observed for EBV-induced DNase from lymphoblastoid cell extracts. In addition, the enzyme was recognized by both immunoglobulin G and A serum fractions from patients with nasopharyngeal carcinoma (NPC). From these results and previous studies which demonstrated the value of antibody titers to this viral DNase as an NPC marker, it appears that EBV-encoded DNase produced in a heterologous expression system could be used in the development of a specific and early NPC diagnosis test.
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PMID:Purification and properties of Epstein-Barr virus DNase expressed in Escherichia coli. 215 38

A novel human papillomavirus type (HPV) was cloned from an invasive cervical carcinoma. The viral clone showed no homology with other known prototypes of HPV (HPV-1 through HPV-57), except HPV-33 by Southern blot analysis under stringent conditions. It showed less than 20% homology to HPV-33 by reassociation kinetic analysis. The restriction endonuclease map of the clone was different from those of other HPV types and its predicted genome organization surmised by hybridization with subgenomic fragment probes of HPV-33 DNA showed the typical HPV genome organization. The results indicate that this clone is a new type of HPV, designated as HPV-58, distinct from the other known types of HPV. HPV-58 was detected in none of 6 specimens of cervical condylomata acuminata, in 7 of 58 specimens of cervical intraepithelial neoplasia, and in 4 of 50 specimens of invasive cervical carcinoma studied in Nagano prefecture, Japan.
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PMID:Molecular cloning of a novel human papillomarvirus (type 58) from an invasive cervical carcinoma. 216 40

Southern blot analysis of 6 human bladder carcinoma cell lines revealed amplification of the epidermal growth factor receptor (EGFR) gene in the KU1 cell line. The amplification of the gene was about 4-fold as compared with that of human placental DNA. Several restriction endonuclease digestions revealed that there was no gross rearrangement of the EGFR gene in KU1. Northern blot analysis showed normal 10 and 5.6 kb of EGFR gene-related mRNA species. 125I-EGF binding revealed 2 distinct EGF binding sites on KU1 cells: high-affinity sites 5.7 X 10(5) receptors per cell with 1.1 nM Kd and low-affinity sites 2.3 X 10(6) receptors per cell with 7.4 nM Kd. The number of the EGFR was compatible with that of the A431 squamous carcinoma cell ine which has an amplified, rearranged and over-expressed EGFR gene. Solid-phase immuno-isolation analysis showed a single 170 kDa EGFR protein in KU1 as well as in A431. Unlike other cell lines with amplified and over-expressed EGFR gene, anchorage-dependent growth of KU1 was stimulated but not inhibited by EGF. Moreover, anchorage-independent growth of KU1 was stimulated by EGF.
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PMID:EGF stimulates anchorage-independent growth of a human bladder carcinoma cell line (KU1) with an amplified and over-expressed EGF receptor gene. 260 69


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