Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2005: ¥1,800,000 (Direct Cost: ¥1,800,000)
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Research Abstract |
Double-strand breaks (DSBs), which are extremely cytotoxic DNA lesions, activate an extensive array of responses that lead to repair of the damage and allow continuation of cellular life. The nuclear protein kinase ATM is regarded as the primary activator of this network, phosphorylating key proteins in numerous signaling pathways. The highly conserved MRN complex plays a role in DSB repair. While NBS1 is a target of ATM, recent observation has shown that the MRN complex itself contributes to the direct activation of ATM. ATR, another protein kinase, has been reported to regulate responses to a broad range of damage, including DSB. Whereas ATM is engaged primarily in DSB repair, several observations suggest that ATR has a critical role in virtually all cellular responses to the arrest of DNA replication forks, which are the DNA structures formed during replication. Pyothorax-associated lymphoma (PAL) is non-Hodgkin's lymphoma that develops from chronic inflammation. Free radicals and o
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xidative stress generated in the inflammatory lesions could cause DNA damage and thus provide a basis for lymphomagenesis. Potential involvement of ATM and ATR genes in PAL lymphomagenesis was examined in eight PAL cell lines and clinical samples from five cases. ATM mutations were detected in five of eight PAL lines. All but one of these mutations affected the PI3KK domain, indicating the loss-of-function mutation of ATM gene. Heterozygous mutations of ATR were found in two of eight lines; one a missense and the other a truncation mutation. ATR mutations were also detected in two of five cases in clinical samples from PAL PAL cells with ATR mutation showed a delay or abrogation in repair for IR-induced DNA DSBs or UV induced DNA single-strand breaks (SSBs), and exhibited a defect in p53 accumulation and failure in cell cycle checkpoint at G1-S phase. These findings showed that mutations of ATR gene result in failure for DNA DSB and SSB repair, suggesting the role of ATM and ATR gene mutations in PAL lymphomagenesis. NK/T-cell lymphoma (NKTCL) and chronic active Epstein-Barr virus infection (CAEBV) are relatively frequent, especially in Asia, and are poor in prognosis. Both diseases are proliferative diseases of NK/T cells that show highly complicated karyotypes, suggesting the involvement of chromosomal instability. The whole coding region of the ATR gene was examined in cell lines derived from NKTCL and CAEBV, as well as tumor samples from patients. ATR alterations were detected in two of eight NKTCL and in one of three CAEBV lines. Most aberrant transcripts observed were deletions resulting from aberrant splicing. ATR alterations were also detected in four of 10 NKTCL clinical samples. Alterations in the ATR gene may result in an abnormal response to DNA DSB and SSB break repair, suggesting a role for ATR gene alterations in NKTCL lymphomagenesis. Mutations of DNA DSB repair genes, ATM, MRE11, RAD50, NBS1 and ATR, together with the presence of microsatellite instability (MSI) were examined in 50 leukemialymphoma cell lines. MSI was detected in 13 (26%) lines. Mutations of intronic mononucleotide repeats in ATM and MRE11 were found in nine and six lines, respectively, whereas mutations of mononucleotide repeats of RAD50 were found in only one line, and none were found in either NBS1 or ATR. Frequencies of ATM and MRE11 mutations were significantly higher in MSI-positive than MSI negative lines. These mutations generated aberrant splicing in both genes. The intensity of the aberrant transcript of ATM (497de122) was stronger in five lines harboring mononucleotide mutations of 2 by or more than in the lines without or with a 1bp mutation. The intensity of the aberrant transcript of MRE11 (315de188) was stronger in four lines with mononucleotide mutations than in lines without. The expression levels of ATM and MRE11 transcripts in MSI-positive lines were significantly higher than those in MSI-negative lines. MSI-positive cell lines showed delay or abrogation of DSB repair. The present study suggests that impairment of the MMR system causes aberrant transcripts in the DSB repair genes ATM and MRE11. This might result in inactivation of the DSB repair system, thus inducing an acceleration of genome instability and accumulation of genetic damage. Less
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