Comparison of rat liver and Walker 256 carcinosarcoma tRNAs

Abstract

The complete nucleotide sequences of both rat liver and Walker 256 mammary carcinosarcoma tRNAAsn reveal that they are identical except for the nucleotide present in the wobble position of the anticodon loop. The rat liver tRNAAsn contains the Q nucleoside, whereas the tumour tRNAAsn contains an unmodified guanosine. The tRNAs from both tissues also show significant quantitative differences in the chromatographic mobilities for isoaccepting species of tRNAAsp, tRNAAsn, tRNAHis and tRNATyr. In addition, chromatographic shifts upon cyanogen bromide treatment and analyses of the alkaline hydrolysates of these tRNAs demonstrate that those of tumour origin contain significantly less Q and Q nucleoside than their normal rat liver counterparts.

Distribution and frequencies of post-transcriptional modifications in tRNAs

“…Other positions in tRNA are much less frequently modified and the functions of these modifications are still largely unknown. Only a few positions have never been found modified in any tRNA sequenced so far (these are positions 5,11,23,24,33,42,43,45,53,59, 62, 63, 73-76 and the majority of positions from the variable loop: 13e, 15-17e, 1e, 3-5e, 21-27e).…”

“…One big challenge in the analysis of tRNA modifications concerns the "degree" of modification one can expect for the hypermodified residues (understood as the fraction of experimentally characterized tRNA sequences that contain this modified residue). Partial character of certain modified bases has been fully documented in some cases, [41][42][43][44][45] but the information can usually be retrieved only from the original publications describing the sequencing data, and it is rarely quantitative. Partial modification was often pointed out when the tRNA to be sequenced originated from cells cultivated in different physiological conditions (e.g., aerobic vs anaerobic, different temperature of growth, availability of intermediate metabolites or cofactors, stress conditions, tumor malignancy etc.).…”

“…Partial modification was often pointed out when the tRNA to be sequenced originated from cells cultivated in different physiological conditions (e.g., aerobic vs anaerobic, different temperature of growth, availability of intermediate metabolites or cofactors, stress conditions, tumor malignancy etc.). 42,[46][47][48][49] Currently, new, more sensitive methodologies used to determine whether a given nucleotide is modified or not, allow for more quantitative detection of such cases, which are now reported to be more frequent than initially anticipated. 8,31,50 It was proposed that such situation may reflect the linkage between tRNA modification processes and regulation of metabolism, e.g.…”

A two-dimensional thin layer chromatographic procedure for the sequential analysis of oligonucleotides employing tritium post-labeling

“…Human placenta tRNA l was isolated to a purity of 1.2 nmoles la per A260 unit as previously described (8,9) and further purified by two additional RPC-5 column chromatographic steps at pH 7.6 (9). Complete digestion of the human placenta tRNAVal with la ribonuclease T1 (Sanko) or ribonuclease A (Sigma), two dimensional resolution of the digestion products on PEI-cellulose, and subsequent elution of the ultraviolet visualized fragments were performed as described earlier for human placenta tRNAPhe (4).…”

A factor in serum and amniotic fluid is a substrate for the tRNA-modifying enzyme tRNA-guanine transferase.

“…This phenomenon is explained by variations in the extent of the Q modifications. The tRNA isoaccepting species that contain Q, [Q+]tRNA, are eluted from RPC-5 and Aminex A-28 columns earlier than are their unmodified counterparts, [Q-]tRNA, in which the queuine position is occupied by guanine (3)(4)(5)(6)(7)(8)(9)(10). The additional positive charge of Q is sufficient to explain this chromatographic difference.…”