[Progress on the study of DNA-dependent protein kinase complex].
Explore the source record for details and available documents.
Animals↗
SEARCH · Search PubMed
Results for “Ku Autoantigen”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
[Expression of DNA-PK in hepato- and cholangio-neoplasms and its significance].
OBJECTIVE: To characterize DNA-PKcs and Ku70 expressions in hepato- and cholangio-neoplastic tissues and the association with the degree of malignancy and invasiveness of the tumors. METHODS: The expression of DNA-PKcs and Ku70 was examined in 47 cases of hepato- or cholangio-neoplasm by immunohistochemistry. RESULTS: Ku70 was expressed in all of the neoplastic tissues examined and with a little variation in levels. The highest expression was observed in adenocarcinomas and adenomas. There was no statistically significant association between Ku70 expression level and the degree of their malignancy extent or invasiveness. In contrast to Ku 70, a wide variation in expression levels of DNA-Pkcs was observed among different types of neoplastic tissues. The highest ratio of positive expressing cells was detected in hepatocellular carcinomas (92.1%), which was significantly higher than that in cholangioadeno carcinomas (65.3%) and biliary cystadenocarcinomas (51.9%). Low or no expression level was detected in papillary adenoma cases. DNA-PKcs expression of invasive adenomas and adeno-carcinomas (61.2%) was significantly higher than that of non-invasive adenomas and adeno-carcinomas (30.4%). There was no expression observed in the normal tissues adjacent to the tumors. CONCLUSION: DNA-PKcs is expressed in hepato- and cholangio-neoplasms and its variable level of expression is associated with the types of the tumor and their degree of malignancy and invasiveness. DNA-PKcs could be recognized as a new biomarker for liver neoplasm.
Adenocarcinoma↗
A comparison of response to cisplatin, radiation and combined treatment for cells deficient in recombination repair pathways.
The responses of cells with mutated DNA repair pathways were compared for cisplatin, radiation and combination treatments. The knockout of the nonhomologous endjoining (NHEJ) pathway resulted in increased radiation sensitivity, but no change in cisplatin response in the mouse cells and increased radiosensitivity but decreased cisplatin sensitivity in chicken cells. The mutation of the homologous recombination repair (HR) pathway through XRCC3 in CHO cells resulted in increased radiation and cisplatin sensitivity and to a lesser extent for the Rad54 knockout in the DT40 chicken cells. The combination treatments of cisplatin and radiation showed that inhibition of the HR repair pathway resulted in super additive effects while the inhibition of the NHEJ pathway in DT40 had no effect. In mouse cells the knockout of the NHEJ pathway resulted in reduced super additivity compared to the parental cell lines. These data show that radiation, cisplatin and combination treatment damage is affected differently by the various DNA repair pathways, which could have a range of effects on combination treatments in tumour cells expressing different levels of DNA repair in the various repair pathways.
Animals↗
Cisplatin sensitization by concurrent mild hyperthermia in parental and mutant cell lines deficient in homologous recombination and non-homologous endjoining repair.
The effect of mild hyperthermia on cisplatin sensitization was examined in two cell line pairs, CHO parental AA8 and irsISF, an XRCC3 mutant (deficient in homologous recombination repair), and mouse parental MEF and knockout Ku80 mutants (deficient in non-homologous endjoining repair). The results showed that mild hyperthermia 40, 41 and 42 degrees C given concurrently with cisplatin treatment caused significant sensitization. The degree of sensitization was comparable for the parental and mutant lines, indicating that these repair pathways were likely not involved in cisplatin thermal sensitization. The shorter concurrent treatments cause a larger sensitization than the longer treatments. The reasons for this are not clear, but thermotolerance may be a factor.
Animals↗
Evaluating the combination of molecular prognostic factors in tumor radiosensitivity in rectal cancer.
BACKGROUND/AIMS: The local recurrence ratio following surgery alone is higher in patients with rectal cancer than in those with colon cancer. Preoperative radiotherapy reduces the rate of local recurrence and improves the chances of survival in patients with resectable advanced rectal carcinoma. Identification of predictive indicators of radiosensitivity is useful in selecting patients best suited for preoperative radiotherapy and thus helps to avoid unnecessary preoperative treatment. We investigated whether the combination of Ku, p53, p21, and p16 predicted tumor radiosensitivity. METHODOLOGY: We studied 96 cases with advanced rectal carcinoma. In preradiation biopsy specimens of tumor samples, all immunoreactive nuclei of cells stained positive for Ku, p53, p21, and p16 were evaluated by immunohistochemistry. The expression of p53, p21 and p16 in more than 5% of tumor cells were defined as positive, whereas both Ku70 and Ku86 protein in more than 70% of such cells were defined as Ku positive. The expression patterns of Ku, p53, p21, and p16 were examined for association with tumor radiosensitivity, which was determined according to the criteria of histopathologic assessment of radiotherapy effects. RESULTS: Univariate analysis showed a correlation between the expression patterns of Ku, p53, p21, and p16 and tumor radiosensitivity, while multivariate analysis showed that the expression pattern of Ku and p16 significantly correlated with tumor radiosensitivity. The combination of Ku and p16, or Ku, p53, p21 and p16 was therefore a good predictive marker for tumor radiosensitivity. CONCLUSIONS: These findings tend to support the hypothesis that the combination of Ku, p53, p21, and p16 expression after radiotherapy can act as a marker for radiosensitivity. Further investigation is needed, as the number of cases in this study was limited.
Adult↗
Prognostic significance of Ku70 protein expression in patients with advanced colorectal cancer.
BACKGROUND/AIMS: Ku protein stimulates DNA repair and signals the damage/stress responses, which may affect apoptosis and cell proliferation. The expression of Ku70 has been reported to be related to cell proliferation in a human gastric cancer cell line. However, in colorectal carcinoma, the clinical significance of Ku70 expression remains unclear. We examined the expression pattern of Ku70 in sporadic colorectal cancer and its relation to clinicopathological parameters and survival. METHODOLOGY: We studied 101 consecutive cases of advanced colorectal carcinoma. In resected specimens, the number of cells stained positive for Ku70 protein was evaluated by immunohistochemistry. RESULTS: The percentage of cells expressing Ku70 in different tumors ranged from 4.2-99.7%. The elevated Ku70 expression group comprised 52 of 101 cases (52%). No significant correlation was seen between Ku expression pattern and the clinical parameters except depth of invasion. pTNM stage, histopathological grade and Ku70 were significant variables for prognosis of survival in the multivariate analysis. CONCLUSIONS: This is the first report correlating reduced survival with elevated expression of Ku protein in colorectal adenocarcinoma. Ku may be a new prognostic marker useful for selecting adjuvant treatment strategies.
Adenocarcinoma↗
[Immunologic tests: Anti-Ku antibodies].
Explore the source record for details and available documents.
Antibodies, Antinuclear↗
Clusterin: the missing link in the calcium-dependent resistance of cancer cells to apoptogenic stimuli.
The resistance of cancer cells to artificially induced apoptosis comprises a major pitfall in contemporary chemotherapy. In recent years, a wide range of molecular mechanisms was revealed that allow cancer cells to overcome apoptosis. In prostate, breast, and colorectal cancers, a protein named clusterin was identified with anti- or proapoptotic activity regulated by calcium homeostasis. Reports so far suggest "two faces" of clusterin activity: the calcium-dependent cellular retention of clusterin positively correlates with cell survival, whereas nuclear translocation of this protein promotes cell death in calcium-deprived cells. Better understanding of the proprieties of clusterin isoforms and the molecular mechanisms which regulate its activity provide the prospect of breaking down cancer cells' resistance to apoptosis. Thus one might expect such dual benefits as overcoming the "immune escape" of neoplastic cells and reducing the doses of cytostatic drugs, with a concomitant reduction in the side effects of chemotherapy.
Animals↗
Evaluation of adaptive responses to cisplatin in normal and mutant cell lines with mutations in recombination repair pathways.
Cell lines mutant in specific DNA repair pathways were used to determine if these pathways are involved in adaptive responses. For these studies, the effect of deficiencies in homologous recombination repair (HR) were studied in the parental AA8 and mutant (irs)ISF cell line pair and for deficiencies in the nonhomologous endjoining (NHEJ) pathway in the mouse MEF parental and Ku80 mutant cell line pair. The results showed that the XRCC3 mutation in the HR-deficient mutant inhibited adaptive responses to low doses of cisplatin and radiation. The parental lines showed transient adaptive responses to both low-dose cisplatin and radiation treatment. For the mouse MEF and the Ku80 cells, no adaptive responses were observed in either cell line. However, there was an initial transient sensitization response followed by partial recovery. Thus, it appears that the HR repair system may be involved in the adaptive response to cisplatin and radiation. For the NHEJ repair system the question could not be answered since no adaptive responses were evident in the parental line.
Animals↗
Altered expression of DNA double-strand repair genes Ku70 and Ku80 in carcinomas of the oral cavity.
The relevance of double-strand DNA repair genes has been demonstrated in several tumours. The main aim of this study was to analyse the expression of the heterodimers Ku70 and Ku80, building regulatory subunits of the DNA-dependent protein kinase in 40 oral carcinomas. Ku70 expression was found in 87.5% of grade 1 and grade 3 tumours and in 82.9% of grade 2 carcinomas. Ku80 presence was noted in 87.5% of grade 1 tumours, 82.9% of grade 2 tumours and in all grade 3 tumours. Ku70-positive cells were present in 90.5% of tumours without and in 80% of tumours with lymphatic metastases. A similar relationship was found for Ku80 expression. Additionally, the expression of Ku70 was highly significantly related to smoking habits. Our results demonstrated that defects of DNA double-strand repair genes play an important role in the tumour progression of oral carcinomas.
Adult↗
[T cell receptor gene recombination in human T-cell leukemia cell line jurkat].
BACKGROUND & OBJECTIVE: Recently, recombination activating gene (RAG)-mediated transposition has been found to contribute to chromosomal translocation and may be related to the occurrence of lymphoid malignancy; however, the underlying mechanism is unclear. Our previous study showed a co-expression of RAG1 and RAG2 in a human T-cell leukemia cell line Jurkat, which represents a mature stage of T cell development, and that the mRNA levels could be regulated by T cell activators. This study was to determine RAGs-mediated T cell receptor(TCR) gene recombination in Jurkat cells, and thus to provide a new thoughtway for studying the correlations of TCR gene recombination to lymphoid malignancy. METHODS: TCR Dbeta-Jbeta signal joint T-cell receptor excision DNA circles (sjTRECs) were determined by nested and semi-nested polymerase chain reaction (PCR). Double-strand DNA breaks at recombination signal sequences (RSSs) in the TCR beta chain locus were detected by ligation-mediated polymerase chain reaction (LM-PCR). TdT and Ku70/Ku80 were detected by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Characteristics of junctional diversity of Dbeta2-Jbeta2 sjTRECs and ds RSS breaks associated with Dbeta2 5'-site and Dbeta2 3'-site were detected in DNA of Jurkat cells. TdT and Ku70/Ku80 were also detected. CONCLUSION: There is an ongoing TCR gene recombination in Jurkat cells. Jurkat may be used as an ideal cell line model for studying the correlation of TCR gene recombination to lymphoid malignancy.
Antigens, Nuclear↗
[Illegitimate recombination as a possible mechanism of DNA-topoisomerase II induced chromosomal rearrangements].
Using semi-quantitative PCR-based approach, we have shown that the breakpoint cluster region of the AML1 gene was associated with the nuclear matrix. We have demonstrated that inhibition of topoisomerase II by etoposide stimulates the appearance of histone gammaH2AX foci, an indicator for the presence of DNA double-strand breaks. Furthermore, the major part of these foci was associated with the nuclear matrix. We also visualized nuclear matrix--associated multiprotein complexes involved in topoisomerase II--induced DNA double-strand break repair. Colocalization studies have demonstrated that these complexes included the principal components of the non-homologous end joining repair system (Ku80, DNA-PKcs and DNA ligase IV). Thus, it is reasonable to suggest that the non-homologous DNA end joining is a possible mechanism of topoisomerase II--induced chromosomal rearrangements.
Antigens, Nuclear↗
cDNA analysis of gene expression associated with DNA-dependent protein kinase activity.
DNA-dependent protein kinase (DNA-PK) is thought to play a pivotal role in DNA double-strand break repair. We recently demonstrated the association of DNA-PK activity in peripheral blood lymphocytes (PBL) with the incidence of chromosomal aberrations and the risk of cancer. In this study, we applied cDNA array technology to find the expression of genes which are associated with DNA-PK activity in PBLs with various levels of DNA-PK activity. Most genes correlated with DNA-PK activity involved cell cycle regulation. Moreover, the transcription factor E2F1, which plays an important role in cell cycle progression, exhibited strong correlation with the DNA-PK activity and Rbp130, which is considered a negative regulator of E2F, showed inverse correlation with DNA-PK activity. In silico promoter analyses showed the presence of at least one E2F binding site in the promoter regions of Ku70, Ku86, DNA-PKcs and genes associated with DNA-PK activity. In order to examine the relationship among the E2F1 expression, the expression of genes related with DNA-PK activity, and DNA-PK activity, we activated PMLs by PHA to progress the cell cycle. After PHA activation of PML, the expression of E2F1 and DNA-PK activity increased. The expression of most genes in PHA-stimulated PBLs had a similar relationship with DNA-PK activity to that without PHA stimulation. These results indicate that the E2F transcription factor may regulate the concerted expression of genes related with DNA-PK activity.
Antigens, Nuclear↗
Ku polypeptides synthesized in vitro assemble into complexes which recognize ends of double-stranded DNA.
The Ku protein is composed of two polypeptide subunits, p70 and p80, and binds DNA ends in vitro. Previous studies suggested that p70 and p80 are physically associated in vivo, although such an association may have been mediated by DNA. We have now utilized full-length Ku polypeptides synthesized in vitro to examine the association of p70, p80, and linear DNA to form a complex. In gel filtration chromatography, p70 migrates as a 70-kDa structure, whereas p80 migrates at 150 kDa. Co-translation of the two cDNAs yields complexes which migrate at 300 kDa and contain equimolar quantities of the p70 and p80 polypeptides, providing direct evidence that p70 and p80 assemble into a complex in the absence of DNA. To demonstrate that this recombinant protein complex binds DNA, we developed a radiolabeled protein electrophoretic mobility shift assay. When radiolabeled proteins synthesized in vitro were incubated with linear DNA and fractionated in a nonreducing, nondenaturing gel, a band representing a complex of p70, p80, and the DNA was seen. Formation of this Ku-DNA complex required free DNA ends, and binding to DNA ends was not observed with individual p70 or p80 subunits. DNA binding was not reconstituted by mixing the individual subunits together. These studies thus demonstrate that it is the complex of p70 and p80, not individual p70 or p80, which possesses the DNA binding properties previously described for native Ku protein. These results provide new information about the assembly, structure, and DNA binding properties of the Ku protein.
Antigens, Nuclear↗
[Clinical significance of anti-Ku antibodies].
Explore the source record for details and available documents.
Antibodies, Antinuclear↗
Antibodies against Ku protein in sera from patients with autoimmune diseases.
Immunoaffinity-purified Ku protein was used to screen sera from patients with systemic lupus erythematosus (SLE), scleroderma, myositis and Sjögren's syndrome for anti-Ku antibodies in a quantitative immunoblot assay. Sixteen percent of the 159 studied sera were reactive with the Ku protein; significantly increased frequencies of anti-Ku antibodies were found in SLE (19%) and scleroderma (14%) sera. Patients with myositis and Sjögren's syndrome showed similar frequencies. All positive sera had antibodies to the 86 kD subunit of Ku protein; only one serum did not react with 70 kD subunit. Frequencies of other autoantibodies were compared in anti-Ku positive and negative patients. Only anti-Sm antibodies, especially in the absence of anti-nRNP, appear to be associated with the presence of anti-Ku antibodies. A strong correlation between anti-Ku antibodies and the class II HLA antigen DQw1 (89% of the positive sera) was observed, suggesting participation of MHC genes in the mounting of the anti-Ku immune response.
Antigens, Nuclear↗
cDNA-derived amino acid sequence of the 86-kDa subunit of the Ku antigen.
The Ku antigen is a DNA-associated nuclear protein recognized by sera from patients with autoimmune diseases. It consists of two polypeptides of 86 and 70 kDa. cDNA clones encoding the 86-kDa subunit of the Ku antigen were isolated by probing lambda gt11 recombinant cDNA expression libraries with a monoclonal antibody specific for this protein. The amino acid sequence deduced from the cDNA comprises 732 amino acids and corresponds to a protein with molecular weight of 81.914. Nineteen residues at the NH2 terminus determined by protein sequencing corresponded to the sequence deduced from the cDNA. The predicted amino acid sequence contains a region with repeating leucine residues similar to the "leucine zipper" structure observed in the c-myc, v-myc, and c-fos oncogene products. The largest cDNA hybridized to 2.7- and 3.4-kilobase poly(A)+ mRNAs from HeLa cells. The cDNA clones expressed fusion proteins immunoreactive with the monoclonal antibody and sera from patients with autoimmune diseases.
Amino Acid Sequence↗
Characterization of the DNA-binding protein antigen Ku recognized by autoantibodies from patients with rheumatic disorders.
We have characterized the biochemical nature of the Ku protein, the antigen recognized by autoantibodies from certain patients with scleroderma-polymyositis overlap syndrome. From extracts of HeLa cells labeled with [32P]orthophosphate, anti-Ku antibodies precipitated a high molecular weight nucleic acid identified as DNA because of sensitivity to DNase I and resistance to RNase. From extracts of cells labeled with [35S] methionine, these antibodies precipitated two polypeptides of 70,000 and 80,000 Da. These proteins were purified using immunoaffinity column chromatography. In immunoblots most sera containing anti-Ku antibodies recognized both Ku proteins but one serum bound only to the 70,000-Da subunit. When nucleosomal segments of chromatin were used as antigen, anti-Ku antibodies precipitated dinucleosomes and larger forms of chromatin but not mononucleosomes. Thus, the Ku antigen is a novel DNA-binding protein that is at least partially exposed on nucleosomal segments of chromatin.
Antigens, Nuclear↗