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Functional Role of Ribosylthymine in Transfer RNA

1980; Wiley; Volume: 104; Issue: 1 Linguagem: Inglês

10.1111/j.1432-1033.1980.tb04396.x

1432-1033

Theo Dingermann, Friedbert PISTEL, Helga Kersten,

Mass Spectrometry Techniques and Applications

In some eucaryotes tRNA species occur containing exclusively uridine or ribosylthymine at position 54. In other eucaryotic tRNA species varying ratios of uridine to ribosylthymine have been found. To elucidate whether variation in the degree of U 54 methylation in eucaryotic tRNA plays a functional role in protein synthesis, polysomal and overal cellular tRNAs in the lower eucaryote Dictyostelium discoideum were characterized during vegetative growth and the early developmental preaggregation stage. The nucleoside composition of the tRNAs was determined and U 54 ‐containing tRNA species were detected after methylation in vitro with the rT‐forming enzyme from Escherichia coli. The following results were obtained. (a) During vegetative growth, 85% of the tRNAs present on polysomes contain rT at position 54, whereas tRNA species containing U 54 accumulate in non‐polysomal cell fractions. (b) Overall cellular and polysomal tRNAs from the preaggregation stage are less modified with respect to rT 54 than those from the vegetative growth stage; in addition pseudo‐uridine (Ψ) and 5‐methylcytidine (m 5 c) are partially lacking at tRNA positions not yet identified. (c) The rT‐containing tRNAs predominate on polysomes also during preaggregation; however, only 65% of the polysomal tRNAs contain ribosylthymine. The results demonstrate that in D. discoideum , the major class tRNAs utilized for protein synthesis includes molecular species in which a U 54 residue has been methylated to rT. The ratio of rT 54 to U 54 in these tRNAs declines at a physiological stage at which the rate of protein synthesis is reduced. Since in vitro an eucaryotic U 54 ‐containing tRNA of this class translates mRNA less efficiently than its rT‐containing counterpart [K. B. Marcu and B. S. Dudock 1976 Nature (Lond.) 261 , 159–162], it is assumed that these U 54 ‐containing tRNAs are rate‐limiting in the elongation of peptide chains in eucaryotes.