Establishment of DNA repair-deficient or-superproficient mice and their development and molecular basis of genetic instability

我们将neo基因插入到小鼠XPA基因的外显子4中，建立了XPA基因敲除小鼠。XPA基因敲除小鼠既没有明显的生理异常，也没有明显的病理改变，但在核苷酸切除修复方面存在缺陷，对UVB或DMBA诱导的皮肤癌非常敏感。我们克隆了一个负责紫外线敏感的啮齿动物突变细胞的基因，该基因属于遗传互补组5(ERCC5)，发现ERCC5也是XPG的致病基因。我们将neo基因插入到小鼠XPG基因的氨基末端，建立了XPG基因敲除小鼠。XPG基因敲除小鼠是以孟德尔方式获得的，但纯合子在断奶时死亡，这表明除了核苷酸切除修复外，XPG基因还发挥着重要作用。我们克隆了小鼠MGMT(O^6-甲基鸟嘌呤DNA甲基转移酶)基因和小鼠MTH1基因(8-oxo-dGTP酶基因)，建立了MGMT或MTH1基因敲除小鼠。MGMT基因敲除小鼠发育几乎正常，但MGMT基因敲除小鼠的体重比正常小鼠轻15%，它们对甲基亚硝脲敏感。纯合子小鼠的造血细胞和免疫细胞明显减少，消化器官的粘膜受到严重侵蚀。MTH1小鼠没有表现出自发性癌症的高发病率。我们克隆了RAD51基因的小鼠同源基因，发现RAD51基因敲除的纯合子在胚胎发生中是致命的。我们建立了HITEC小鼠，利用它我们可以检测到小鼠rpsL转基因的体细胞突变。结果发现，经甲基亚硝脲处理后，HITEC小鼠脾、胸腺、肺和肝脏rpsL基因突变频率显著增加。几乎所有的突变都是从G到A的转变。

We established the XPA (xeroderma pigmentosum group A) knockout mice by insertion of neo gene into exon 4 of the mouse XPA gene. XPA knockout mice showed neither obvious physical abnormalities nor pathological alterations, but were defective in nucleotide excision repair and highly susceptible to UVB- or DMBA-induced skin carcinogenesis. We cloned a gene responsible for the UV-sensitive rodent mutant cells which belong to the genetic complementation group 5 (ERCC5) and found that the ERCC5 was also a causative gene for XPG.We established XPG knockout mice by insertion of neo gene into the amino terminal region of the mouse XPG gene. The XPG knockout mice were obtained in a Mendelian fashion, but the homozygotes died by the time of weaning, indicating that the XPG gene play an essential role besides nucleotide excision repair. We cloned mouse MGMT (O^6-methyl guanine DNA methyltransferase) gene and mouse MTH1 gene (8-oxo-dGTPase gene) and established MGMT- or MTH1-knockout mice. MGMT knockout mice developed almost normally but the body weight of the MGMT knockout mice was 15% smaller than normal littermates and they were hypersensitive to methylnitrosourea. The homozygotes developed significant decrease of hematopoietic and immune cells and severe erosion of the mucous membrane of the digestive organs. The MTH1 mice did not show high incidence of spontaneous cancers. We cloned mouse homologue of the Rad51 gene and found that the Rad51 knockout homozygotes are lethal in the embryogenesis. We established HITEC mice with which we can detect somatic mutations in the rpsL transgene in the mice. We found that the mutation frequency in the rpsL gene of the spleen, thymus, lung and liver of the HITEC mice was significantly increased after treatment of the mice with methylnitrosourea. Almost all the mutations were G to A transitions.