The biosynthesis of transfer ribonucleic acid in the developing rat brain and in cultured glial cells

The biosynthesis of tRNA was investigated in cultured astroglial cells and the 3-day-old rat brain in vivo. In the culture system astrocytes were grown for 19 days and were then exposed to [3H]guanosine for 1.5-7.5 h; 3-day-old rats were injected with [3H]guanosine and were killed 5-45 min later. [3H]tRNA was extracted, partially purified, and hydrolyzed to yield [3H]guanine and [3H]methyl guanines. The latter were separated from the former by high performance liquid chromatography and their radioactivity determined as a function of the time of exposure to [3H]guanosine. The findings indicate that labeling of astrocyte tRNA continued for 7.5 h and was maximal, relative to total RNA labeling, at 3 h, while in the immature brain tRNAs were maximally labeled at 20 min after [3H]guanosine administration. The labeling pattern of the individual methyl guanines differed considerably betweren astrocyte and brain tRNAs. Thus, [3H]1-methylguanine represented up to 35% of the total [3H]methyl guanine radioactivity in astrocyte [3H]tRNA, while it became only negligibly labeled in brain [3H]tRNA. Conversely, brain [3H]tRNA contained more [3H]N2-methylguanine than did astrocyte [3H]tRNA. Approximately equal proportions of [3H]7-methylguanine were found in the [3H]tRNAs of both neural systems. The [3H]methylguanine composition of brain [3H]tRNA was followed through several stages of tRNA purification, including benzoylated DEAE-cellulose and reverse phase chromatography (RPC-5), and differences were found between the [3H]methylguanine composition of RPC-5 fractions containing, respectively, tRNAlys and tRNAphe. The overall results of this study suggest that developing brain cells biosynthesize their particular complement of tRNAs actively and in a cell-specific manner, as attested by the significant differences in the labeling rates of their methylated guanines. The notion is advanced that cell-specific tRNA modifications may be a prerequisite for the successful synthesis of cell-specific neural proteins.