Abstract
Most of the oxidized bases generated in mammalian genome under a physiological condition are repaired by base excision repair (BER) pathways. DNA glycosylases remove altered bases and form apurinic/apyrimidinic sites at the first step of BER. Mouse endonuclease VIII like glycosylase 1 (mNEIL1) is a DNA glycosylase against various oxidized pyrimidines, and also plays a key role during DNA replication. Two types of mouse variant mRNAs, NCBI AAH43297 (variant1) and NCBI BAC30707 (variant2), are detected in mammary gland tumors and in aorta/vein, respectively. Interestingly, there is no report about such variants in human tissues. To study significance of these variants, we prepared recombinant proteins of mNEIL1and these variants using the pET22b expression system. When we examined the DNA glycosylase activities against thymine glycol, a typical oxidized damage, no activity was detected. With polyclonal antibody developed against recombinant mNEIL1, the variants as well as NEIL1 were detected in liver extract. To study the expression in mouse tissues, we prepared primer sets to amplify specifically the cDNAs of mNEIL1 or variants and performed RT-PCR using total RNAs prepared from Jcl:ICR mouse kidney and liver. Since apparently more abundant expressions of variant mRNAs were observed in kidney, suggesting that there are differences among the mRNA levels in normal mouse organs. The analyses of the mRNA levels in various mouse organs and the enzymatic activities against a series of altered pyrimidines are in progress.
