@misc{oai:repo.qst.go.jp:00069487,
 author = {Morohoshi, Fumiko and Tomita, Masanori and Hashimoto, Mitsumasa and Otsuka, Kensuke and Hayata, Isamu and Iwabuchi, Kuniyoshi and Tauchi, Hiroshi and Sakai, Kazuo and et.al and 冨田 雅典 and 橋本 光正 and 大塚 健介 and 早田 勇 and 田内 広 and 酒井 一夫},
 month = {Jul},
 note = {The BRCT domain is an evolutionarily conserved domain found in a large number of proteins involved in DNA repair, recombination and damage checkpoints. To elucidate the function of the PTIP protein which carries 6 BRCT domains, we constructed constitutive (chPTIP-/-/-) and conditional PTIP-null mutants (tet-off hPTIP, chPTIP-/-/-) in chicken DT40 cells. In contrast to mouse PTIP-null mutant cells. DT40 PTIP-null mutants are viable, but appear to have a defective proliferative capacity, since a high proportion of dead cells appear during culture growth and colony formation ability in methyl cellulose containing medium was less than 1% of that of wild type cells. Conditional PTIP mutants express human PTIP (hPTIP) in the absence of doxycycline (Dox) but expression levels decrease to almost undetectable levels within 48 hours after the addition of Dox. Starting from the 7th day after the addition of Dox, the cell culture s begin to produce deal cells and show chromatid-type gaps. Conditional PTIP mutants become sensitive to the lethal effects of ionizing radiation (IR) at 1 to 2 days after the addition of Dox [D10(+Dox)/D10(-Dox)=0.8], but not to the effects of UV and MMS, and the cells display low levels of resistance to MMC. The hPTIP polypeptide consisting of the two N-terminal BRCT domains was able to extensively complement cell proliferation defects and high radiation sensitivity of constitutive PTIP mutants.
Since PTIP mutants showed a higher sensitivity to IR, we examined whether they were defective in double strand break repair or damage induced checkpoints. Western blot analysis revealed that the phosphorylation kinetics of H2AX after IR irradiation was similar in PTIP mutants and in wild type cells. Rad51 foci formation and disappearance after IR irradiation was normal, and homologous recombination operated normally in PTIP mutants. These results indicate that the PTIP protein is not involved in double strand break repair. The G2 block after IR irradiation also operated normally in PTIP mutants. However IR-induced phosphorylation of S345 in Chk1 was higher in PTIP mutants than in wild type cells. This enhanced phosphorylation of Chk1 S345 was not observed after UV irradiation in PTIP mutants. These results suggested that repair of IR induced DNA damage or IR induced signaling was defective in PTIP mutants., 13th International Congress of Radiation Research},
 title = {Chicken DT40 PTIP-null mutants are viable, but defective in proliferation and highly sensitive to ionizing radiation.},
 year = {2007}
}