Effects of post-transcriptional base modifications on the site-specific function of transfer RNA in eukaryote translation.

The site-specific function in translation of several naturally occurring mammalian transfer RNAs has been studied in a series of investigations with some similarities to studies in other laboratories of tRNAs in suppression. Equal amounts of aminoacyl-tRNA isoacceptors with contrasting isotopes were added in pairs to reticulocyte lysates and allowed to incorporate their amino acids into rabbit globin. Rates of incorporation from unlimiting amounts of each isoacceptor into the corresponding amino-acid-containing sites were determined. The tRNAs of each isoacceptor pair differed as to post-transcriptional base modifications. The natural occurrence of these isoacceptors can be correlated with rates of cellular division, with more rapidly dividing and neoplastic cells containing hypomodified tRNA. The overall incorporation of lysine into globin from a fully modified tRNALys that decodes AAG is faster by 25 to 30% than from the corresponding hypomodified tRNALys. There is considerable scatter in values for incorporation ratios at different lysine-containing sites, with the hypomodified isoacceptor even being preferred at one site. The AAG decoding isoacceptors are capable of translating AAA although much more slowly than AAG. In translating AAA, in contrast to translating AAG, the hypomodified tRNALys isoacceptor is preferred. A Y base-deficient hypomodified tRNAPhe isoacceptor found only in some kinds of rapidly dividing tumor cells donates its phenylalanine preferentially to globin in competition with the fully modified Y-containing tRNAPhe of liver by 15 to 17%. There is a considerable range of incorporation ratios at the different phenylalanine-containing sites of the globin subunits. No correlation can be made between the isoacceptor preferred and the phenylalanine codon being translated. The incorporation of histidine from a fully modified tRNAHis-containing Q base in its anticodon, compared with that from the hypomodified counterpart isoacceptor that lacks Q base and that occurs in rapidly dividing cells, showed no difference in their ability to incorporate overall or into individual histidine-containing sites. There is little evidence that adjacent bases or codons in messenger RNA affect the tRNAs preferred in the translation of most sites. A striking pattern of tRNA preference was observed in three cases in which there are tandem codons, with the same codon appearing twice in succession.(ABSTRACT TRUNCATED AT 400 WORDS)