Studies on the function of the non-primer tRNAs associated with the 70 S RNA of avian myeloblastosis virus.

Significant amounts of three tRNAs are associated with the 70 S RNA of avian myeloblastosis virus (AMV). The temperatures at which they are half dissociated from the 70 S RNA in 50 mM NaCl and their respective quantities relative to 35 S RNA are: tRNAArg, 51 degree C, 1.6; tRNALys, 57 degree C, 0.7 and tRNATrp, 76 degree C, 1.0. Possible functions for the non-primer tRNAs (tRNAArg and tRNALys) were evaluated by determining the effect of their thermal dissociation on: (a) conversion of 70 S to 35 S RNA, (b) capacity of 70 S and/or 35 S RNA to be translated in vitro, and (c) capacity of 70 S and/or 35 S RNA to be reverse transcribed in vitro. Conversion of 70 S to 35 S RNA occurred with a tm of 56 degree C and is consistent with the hypothesis that tRNALys might be involved in joining two 35 S RNA subunits to form the 70 S RNA complex. There was no indication that the association of either tRNAArg or tRNALys influenced the rate or quality of translation of 70 S or 35 S RNA. A decrease in the rate at which 70 S RNA is transcribed occurs in parallel with the dissociation of tRNAArg and tRNALys.

Efficiency and fidelity of cell-free protein synthesis by transfer RNA from aged mice.

Transfer RNAs (tRNAs) from heart, kidney, liver, and spleen of mature (10-12 months old) and aged (29 months old) C57BL/6 mice were tested for their ability to translate encephalomyocarditis viral RNA in a tRNA-dependent cell-free system derived from mouse ascites tumor cells. The rates of in vitro protein synthesis were compared as a function of tRNA concentration, and the fidelity of translation was examined by sodium dodecyl sulfate gel electrophoresis and isoelectric focusing of the viral polypeptides synthesized in vitro. No significant age-related differences in either the efficiency or fidelity of synthesis were discovered, indicating that alterations in tRNAs are probably not involved in the cellular aging of these tissues.

Alteration of the Q family of transfer RNAs in adult Drosophila melanogaster as a function of age, nutrition, and genotype.

Four tRNAs are known to contain Q, a hypermodified form of guanosine, in one of their isoacceptor forms; these are tRNATyr, tRNAHis, tRNAAsp, and tRNAAsn. The first three have been examined in Drosophila melanogaster at different ages in four genotypes. As the adult flies age, the ratio of the Q/non-Q isoacceptors increases, and the rate of increase is more rapid for Samarkand than for Oregon-R (two wild-type strains). Similarly, two other strains that carry the mutations su(s)2 v; bw and v; bw also differ in the rate of isoacceptor alteration. Diet has a marked effect on the ratio of Q/non-Q isoacceptors for each tRNA. tRNALeu does not change with age or diet and is believed not to contain Q in any of its isoacceptors. The possible role of Q in altering the rate of protein synthesis was examined in a cell-free, tRNA-dependent, mRNA-dependent system. No essential effect on the rate or extent of protein synthesis was detected upon comparison of Q-enriched or Q-deficient tRNA. The ratio of Q/non-Q isoacceptors for these tRNAs is clearly controlled by age-related and diet-related mechanisms that can modulate the amount of Q in the tRNA. The relationship of these studies to Strehler's hypothesis of age-related control of protein synthesis is discussed.

Undermethylated transfer RNA does not support phage RNA-directed in vitro protein synthesis.

A cell-free protein synthesis system from Escherichia coli Q13 was depleted of mRNA and tRNA so that restoration of maximum activity was dependent of the addition of these components. Protein synthesis, directed by either MS2 or Qbeta phage RNA, was stimulated significantly by the addition of normal tRNA from either E. coli Q13 or B. In contrast, undermethylated tRNA from methionine-starved E. coli RCrel did not cause this stimulation. It is concluded that undermethylated tRNA Lacks sufficient base modifications to function in protein synthesis.