An Unusual Transfer RNA (Guanine-2-)-Methyltransferase from Transplantable Rat Mammary Tumors

We have previously demonstrated that (guanine-2-)-methyltransferase activity in extracts from 9,10-dimethyl-1,2-benzanthracene-induced rat mammary tumors differs from that of nonneoplastic mammary tissue. In this report, we explore further the nature of these differences by purification and characterization of the two major transfer RNA (tRNA) (guanine-2-)-methyltransferases from transplantable mammary tumors and proliferating mammary glands from pregnant rats. The position 10-specific (guanine-2-)-methyltransferases (2mGI) from proliferating rat mammary gland and mammary tumor were found to have similar properties with respect to molecular weight, substrate specificity, and elution behavior on ion-exchange columns. In addition, no tissue-specific differences were observed when the mammary tumor and mammary gland 2mGI activities were compared with those of purified rat liver enzyme. In contrast, the position 26-specific (guanine-2-)-methyltransferase (2mGII) from mammary tumors was seen to possess properties different from both the nontumorous mammary gland and liver enzyme. The tumor 2mGII activity showed unusual elution behavior on diethylaminoethyl-Sephadex, eluting along with the 2mGI activity. A small difference in molecular weight was detected between tumor and nontumorous 2mGII activities. Examination of the tumor enzyme in comparison with the well-characterized 2mGII from rat liver indicated that the mammary tumor 2mGII methylated a broader range of tRNA substrates. In particular, mature yeast phenylalanine-specific tRNA, which is methylated in vivo at all major eukaryotic methylation sites and should not be a substrate for eukaryotic methylating enzymes in vitro , could be methylated at low levels by the tumor enzyme. Two-dimensional electrophoretic fingerprint maps of T1 RNase-digested phenylalanine-specific tRNA from Escherichia coli methylated in vitro showed the presence of a methylated oligonucleotide which could not be correlated with normal sites of methylation on the tRNA. From these results, it appears that the mammary tumor 2mGII can methylate at some unusual site(s) on the tRNA molecule.