Morland Ingrid, Rolseth Veslemøy, Luna Luisa, Rognes Torbjørn, Bjørås Magnar, Seeberg Erling
Department of Molecular Biology, Institute of Medical Microbiology, University of Oslo, Rikshospitalet, 0027 Oslo, Norway.
Nucleic Acids Res. 2002 Nov 15;30(22):4926-36. doi: 10.1093/nar/gkf618.
The mild phenotype associated with targeted disruption of the mouse OGG1 and NTH1 genes has been attributed to the existence of back-up activities and/or alternative pathways for the removal of oxidised DNA bases. We have characterised two new genes in human cells that encode DNA glycosylases, homologous to the bacterial Fpg (MutM)/Nei class of enzymes, capable of removing lesions that are substrates for both hOGG1 and hNTH1. One gene, designated HFPG1, showed ubiquitous expression in all tissues examined whereas the second gene, HFPG2, was only expressed at detectable levels in the thymus and testis. Transient transfections of HeLa cells with fusions of the cDNAs to EGFP revealed intracellular sorting to the nucleus with accumulation in the nucleoli for hFPG1, while hFPG2 co-localised with the 30 kDa subunit of RPA. hFPG1 was purified and shown to act on DNA substrates containing 8-oxoguanine, 5-hydroxycytosine and abasic sites. Removal of 8-oxoguanine, but not cleavage at abasic sites, was opposite base-dependent, with 8-oxoG:C being the preferred substrate and negligible activity towards 8-oxoG:A. It thus appears that hFPG1 has properties similar to mammalian OGG1 in preventing mutations arising from misincorporation of A across 8-oxoG and could function as a back-up repair activity for OGG1 in ogg1(-/-) mice.
与小鼠OGG1和NTH1基因靶向破坏相关的轻度表型归因于存在用于去除氧化DNA碱基的备用活性和/或替代途径。我们在人类细胞中鉴定了两个新基因，它们编码与细菌Fpg（MutM）/Nei类酶同源的DNA糖基化酶，能够去除hOGG1和hNTH1的底物损伤。一个基因命名为HFPG1，在所检测的所有组织中均呈现普遍表达，而另一个基因HFPG2仅在胸腺和睾丸中以可检测水平表达。用cDNA与EGFP的融合体对HeLa细胞进行瞬时转染，结果显示hFPG1在细胞内定位于细胞核并在核仁中积累，而hFPG2与RPA的30 kDa亚基共定位。hFPG1被纯化，并显示其作用于含有8-氧代鸟嘌呤、5-羟基胞嘧啶和无碱基位点的DNA底物。8-氧代鸟嘌呤的去除，而非无碱基位点的切割，呈现碱基依赖性，8-氧代鸟嘌呤：胞嘧啶是首选底物，对8-氧代鸟嘌呤：腺嘌呤的活性可忽略不计。因此，hFPG1在防止因腺嘌呤错掺入8-氧代鸟嘌呤而产生的突变方面具有与哺乳动物OGG1相似的特性，并且在ogg1（-/-）小鼠中可作为OGG1的备用修复活性发挥作用。
