tRNA Methylase Research Articles

Bases methylated in vitro by cell-free extracts of adenovirus-18-induced tumours.

The tRNA methylase activity in vitro of adenovirus-18-induced tumour was studied. The activity expressed per mg of protein was the same for extracts of tumours induced by either adenovirus-12 or adenovirus-18. The methylated bases isolated after incubation with extract from adenovirus-18-induced tumour were N(1)-methylguanine, 1-methyladenine, 3-methyladenine and N(6)-methyladenine.

Specificities of tRNA methylases from mouse liver and plasmocytoma

In vitro methylation of some Escherichia coli specific tRNAs by extracts from mouse normal liver and MOPC 173 plasmocytoma has been studied. The determination of the overall extent of methylation as well as of the repartition of methylated products reveals that tRNA methylases from these two sources have distinct specificities. In particular, plasmocytoma extracts seem to contain one methylase that does not exist in liver extract, the activity of which appears when tRNA Met f is used as a substrate.

Studies of the ethylation of rat liver transfer ribonucleic acid after administration of L-ethionine.

1. The ethylated nucleosides present in tRNA isolated from the livers of rats treated with 0.5g of l-ethionine/kg body wt. were investigated. Evidence that this tRNA contained N(2)-ethylguanine, N(2)N(2)-diethylguanine, N(2)-ethyl-N(2)-methylguanine, 7-ethylguanine, two ethylated pyrimidines and ethylated ribose groups was obtained. 2. Ethylation of bacterial tRNA was catalysed by extracts containing tRNA methylases prepared from rat liver by using S-adenosyl-l-ethionine as an ethyl donor, but the rate of ethylation was 20 times less than the rate of methylation with S-adenosyl-l-methionine as a methyl donor. 3. The principal product of such ethylation in vitro was N(2)-ethylguanine and traces of the other ethylated guanines and pyrimidines found in tRNA isolated from rats treated with ethionine in vivo were also found. 1-Ethyladenine was not formed, although 1-methyl-adenine is a major product of methylation of bacterial tRNA by these extracts, and 1-ethyladenine was not present in the rat liver tRNA isolated from ethionine-treated animals. 4. After injection of actinomycin D (15mg/kg body wt.) or l-methionine (1.0g/kg body wt.) before the ethionine, ethylation of tRNA was diminished by about 80% but not completely abolished. Administration of 1-aminocyclopentanecarboxylic acid (2.5g/kg body wt.) to inhibit the formation of S-adenosyl-l-ethionine inhibited ethylation of tRNA by 44%. 5. These results suggest that not all of the ethylation of tRNA that occurs in the livers of rats treated with ethionine is mediated by the action of tRNA methylases acting with S-adenosyl-l-ethionine as a substrate, but that this pathway does occur and accounts for a major part of the observed ethylation. 6. The results are discussed with reference to ethionine-induced hepatocarcinogenesis.

Nicotinamide and Its Methyltransferase as Apparent Inhibitors of tRNA Methylase

Nicotinamide and Its Methyltransferase as Apparent Inhibitors of tRNA Methylase

Transfer ribonucleic acid methylases of bone. Studies on vitamin A and D deficiency.

Methods were devised for the assay of tRNA methylases of rat bone. The activities of bone tRNA methylases are similar to those from other mammalian tissues. However, unlike reports on liver methylases, no inhibitors were found in the supernatant fraction from pH5 precipitate of bone extracts. The effects of vitamins A and D on the methylation of tRNA by cell-free extracts of rat bone were studied. Deficiency of either vitamin resulted in a decrease in the rate and extent of tRNA methylation, whereas the administration of vitamin A to hypovitaminotic-A rats and vitamin D to hypovitaminotic-D rats increased the rate and extent of tRNA methylation. These effects appear to be apart from changes in ribonuclease activity or in concentrations of calcium or magnesium. No evidence of inhibitors of tRNA methylases was found in bone extracts from vitamin-deficient rats nor of activators in bone extracts from deficient rats given vitamin A or D. The pattern of tRNA methylation under conditions of vitamin A or D deficiency was not changed, suggesting a generalized cellular deficiency. It was of significance to find that the specificity for methylation of specific bases in tRNA was different after the administration of vitamin A as contrasted with the effects of vitamin D. The possible significance of tRNA methylation to the biochemical action of the vitamins on bone is discussed.

Atypical Transfer RNA's And Their Origin In Neoplastic Cells

Publisher Summary This chapter focuses on the structure and synthesis of transfer RNA (tRNA) and discusses the biochemistry of tRNA methylases. Translation is the most complex molecular mechanism known in the living cell. It requires messenger RNA, which is the simplest of the structures involved, even though it is laden with information. The ribosomes are complex structures composed of RNA and at least a score of different proteins. Transfer RNA is the most complex biomacromolecule known. In addition to these major components, there are ancillary ones— enzymes that transfer amino acids to the transfer RNA. Moreover, there are at least 8 soluble protein factors needed for the completion of the synthesis of a protein. Any or all of these numerous components could be a regulatory factor in protein synthesis. There are three different lines of biological evidence that point to transfer RNA as a regulatory factor. Transfer RNA is a pivotal molecule in protein synthesis. It is the link between the amino acids and the message-bearing nucleic acids. The tRNA methylases are a complex family of enzymes that modify the structure of preformed tRNA by the insertion of methyl groups into specific positions in the four main bases of tRNA. The enzymes are species-specific, organ-specific, base-specific, and even site-specific for particular bases.

The inhibition of tRNA methylase activity by nicotinamide and a non-dialyzable inhibitor

A non-dialyzable tRNA methylase inhibitor was isolated from normal adult rat liver. The inhibition caused by this factor was eliminated on prolonged dialysis. However, when nicotinamide was added back to the dialyzed fraction, the inhibitory activity was reconstituted.

Effect of the carcinogen N-nitrosomethylurea on the tRNA methylase activity of cells cultured in vitro.

AbstractA comparison has been made between the activity of enzymes responsible for methylation of tRNA in a permanent line of adult Chinese hamster ovary fibroblasts maintained in in vitro culture, and that of cells derived from this line after exposure in vitro to the alkylating carcinogen N‐nitrosomethylurea. A significantly elevated activity has been observed in several clones of cells which had received a single acute dose of the nitrosamide. A series of chronic treatments in which the single acute dose was given over a period of 8 weeks illustrated the cumulative effect of the fractionated doses on the methylase enzymes, and confirmed the repeatability of findings after a single acute dose. Evidence has been obtained from morphological criteria that a correlation appears to exist between the extent of morphological conversion within the cell population and the activity of the enzymes. Kinetic evidence suggests that this alteration in activity is not related to changes in growth rate alone. These results are discussed in relation to the concept that methylation of tRNA may relate to the degree of maturation and differentiation in a particular cell population.

Transfer ribonucleic acid methylases during the germination of Neurospora crassa.

Transfer ribonucleic acid (tRNA) methylases were studied during the germination of spores in Neurospora crassa. The total methylase capacity and base specific tRNA methylase activities were determined in extracts from cells harvested at various stages of germination. Germinated conidia have a 65% higher methylase capacity than ungerminated conidia. Three predominant methylase activities were found in the extracts, and the relative amount of each activity was different at the various stages. Enzymes from vegetative cells catalyzed significant hypermethylation of tRNA from conidia, whereas conidial enzymes were much less active on tRNA from vegetative cells. The results indicate differences in the tRNA methylase content and tRNA species of conidia and vegetative cells.

Studies on inhibitors of mammalian tRNA methylases

FEBS LettersVolume 16, Issue 1 p. 13-16 Full-length articleFree Access Studies on inhibitors of mammalian tRNA methylases A.E. Pegg, A.E. Pegg Courtauld Institute of Biochemistry, Middlesex Hospital Medical School, London W1P 5PR, EnglandSearch for more papers by this author A.E. Pegg, A.E. Pegg Courtauld Institute of Biochemistry, Middlesex Hospital Medical School, London W1P 5PR, EnglandSearch for more papers by this author First published: July 15, 1971 https://doi.org/10.1016/0014-5793(71)80672-5Citations: 42AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Citing Literature Volume16, Issue1July 15, 1971Pages 13-16 ReferencesRelatedInformation

Transfer RNA methylases from rat liver nuclei

The intracellular localization of transfer RNA methylases has been investigated. In contrast to previous findings with nuclei from aqueous rat liver homogenates, non-aqueous tissue fractionation results in nuclei with a methylating activity 30 times higher than the corresponding cytoplasm. During fractionation, methylating activity rises parallel to the DNA content of the fraction. Methylases from non-aqueous nuclei and those known from the supernatants of aqueous homogenates have the same substrate specificity. It is concluded that the bulk of tRNA methylases is located within the nucleus of the cell.

Transfer RNA methylases during sea urchin embryogenesis

Methylation of tRNA in sea urchin Strongylocentrotus purpuratus, during embryogenesis was studied by labeling with [ Me- 3H]methionine. The changes in methylated bases in tRNA following fertilization were not uniform. The N 2-monomethylguanine was reduced to half and methylated uridylic acids increased 3-fold. Extracts of eggs and of embryos of various stages after fertilization were active in incorporating methyl groups from S- adenosyl- l -[Me- 14 C]methionine into Escherichia coli B tRNA.

A study of tRNA methylases by the dissected molecule method

A study of tRNA methylases by the dissected molecule method

Fragmented [formula omitted] methionine tRNA f as methyl acceptor for rat liver tRNA methylase: Alteration of the site of methylation by the conformational change of tRNA structure resulting from fragmentation

In vitro methylation of E. coli tRNA fMet fragments or their reconstituted molecule by rat liver methylase showed that (1) the whole tRNA molecule was not necessary for recognition by tRNA methylase, and (2) the extent and the site of methylation of fragments differed depending upon the type of fragment used. This indicates that the conformational structure of the methyl acceptor molecule is important for recognition by the tRNA methylase.

The Transfer RNA Methylases of Human Lymphocytes. II. Delayed Induction by PHA in Lymphocytes From Patients With Chronic Lymphocytic Leukemia

Abstract Treatment of lymphocytes from three patients with chronic lymphocytic leukemia (CLL) with phytohemagglutinin (PHA) resulted in a fivefold increase in transfer RNA (tRNA) methylase activity. Evidence was obtained which suggests that the increase included enzyme species not present prior to the induction. As with normal lymphocytes, the time of induction coincided with the formation of fully transformed PHA cells, which is markedly delayed compared with that occurring in normal lymphocytes stimulated with PHA. Although all three patients had an identical pattern of tRNA methylase induction, initial and induced absolute values were higher in the cells of one patient who had 10 per cent immature peripheral blood lymphocytes. The levels of transfer RNA methylase activity were similar in normal and CLL lymphocytes. These data suggest that PHA induces quantitative and qualitative changes in tRNA methylase enzymes of CLL lymphocytes similar to those previously reported in normal lymphocytes but that the sequence of events of PHA interaction with CLL lymphocytes leading to the subsequent enzyme induction is delayed.

The Transfer RNA Methylases of Human Lymphocytes. I. Induction by PHA in Normal Lymphocytes

Abstract Assays were performed that measured both the rate and extent (or total capacity) of transfer RNA methylases in extracts of lymphocytes cultured in the presence and absence of phytohemagglutinin (PHA). The tRNA methylases of human peripheral blood lymphocytes undergoing blastogenesis in culture with PHA had a five- to sixfold increase in rate and a three- to sevenfold increase in extent of methylation of heterologous tRNA. These data suggest that PHA transformed lymphocytes not only contain elevated levels of tRNA methylases, but that the increase includes qualitatively different enzymes from those found in normal peripheral blood lymphocytes. Experiments in which lymphocytes were incubated for various times with PHA revealed that tRNA methylase induction occurred late in or after DNA synthesis and after morphologic transformation, but prior to mitosis. Rate and extent of tRNA methylation increased simultaneously. PHA induction of tRNA methylase activity was dependent on the synthesis of new RNA in lymphocytes cultured from 40 to 45 hours. The increase was not due to different levels of inhibitors or activators or preferential degradation of reaction components. The data suggest that quantitative and qualitative changes occur in the tRNA methylases of the normal human peripheral blood lymphocyte stimulated by PHA to undergo transition to an undifferentiated cell "in cycle." The possible significance of these findings to control of protein synthesis in PHA transformed lymphocytes is discussed.

Transfer RNA methylase alterations in polyoma transformed rat embryo culture cells.

A polyoma-transformed cell line of proven tumorigenicity is demonstrated to have greater tRNA methylase activity than its nontumorigenic parent cell line when cultured, harvested, and assayed under controlled conditions. The marked differences in capacity to methylate heterologous tRNA substrates suggests, but does not prove, that qualitative differences may exist between the methylases of the paired cell lines.

Assay of tRNA methylase enzymes in studies of malignant transformation [formula omitted

Using a standard line of hamster kidney cells cultured in vitro , the activities of specific enzymes concerned with the methylation of transfer RNA has been investigated. Results obtained confirm the observations made in vivo that enzyme preparations from malignant cells have an elevated potential to methylate an exogenous substrate when compared with morphologically normal cells. The similarity between the results obtained with the cultured cells and the neonatal tissue from which the cells were derived confirm the application of this assay procedure to cells grown in vitro . Evidence is presented that the elevated enzyme activity is not solely a function of growth rate but appears to be related to the neoplastic state of the cells. For interpretation of biochemical changes relating to initiation of neoplastic conversion in vitro culture of cells in parallel under identical conditions of growth offers a unique advantage.

Absence of a natural inhibitor of the tRNA methylases from fetal and tumor tissues.

The inhibitor of tRNA methylases of normal adult tissues has been resolved into two fractions, a high molecular weight protein and a low molecular weight (<700) component. The high molecular weight component is absent from embryonic and tumor tissues, both of which are known to possess high tRNA methylase capacity.

Nicotinamide: A natural inhibitor of tRNA methylase

A dialyzable tRNA methylase inhibitor has been isolated from rat liver. By chromatographic and electrophoretic data as well as ultraviolet and mass spectral data, the inhibitor is identified as nicotinamide. Kinetic evidence indicates that nicotinamide is an inhibitor of tRNA methylase.