Modulation of mammalian cell proliferation by a modified tRNA base of bacterial origin

Langgut, Werner; Reisser, Thomas; Nishimura, Susumu; Kersten, H.

doi:10.1016/0014-5793(93)81627-c

Cited by 24 publications

(27 citation statements)

References 24 publications

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“…q stimulates the proliferation of non-transformed NIH-3T3 cells, but inhibits the proliferation of rus-transformed NIH-3T3 cells. Transformation of NIH-3T3 cells with the rus oncogene causes a shift of the metabolism from respiration towards glycolysis even under aerobic conditions as indicated by an increase in lactate production [22]. Cell proliferation, glucose metabolism and the stress response are processes that are controlled by receptors for growth factors or hormones [53-551.…”

Section: Discussionmentioning

confidence: 99%

The nutrient factor queuine protects HeLa cells from hypoxic stress and improves metabolic adaptation to oxygen availability

Reisser

Langgut

Kersten

1994

European Journal of Biochemistry

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Queuine (q), a cyclopentendiol derivative of 7-aminomethyl-7-deazaguanine, is a nutrient factor for lower and higher eukaryotes, except yeast; it is synthesized in eubacteria partly at the level of tRNA. In eukaryotes q is preferentially inserted into the wobble position of specific tRNAs in differentiated and adult tissues, but occurs mainly free in embryonic and fast proliferating cells. HeLa cells grow to a higher cell densitiy under aerobic than under hypoxic conditions only when supplemented with q. Here we show that in hypoxically grown HeLa cells, sufficiently supplied with q, free q accumulated when serum factors become limiting while the respective tRNAs remained completely q deficient. In these cells the levels of lactate dehydrogenase A (LDH A) mRNA and of LDH A protein were at least twofold higher than in aerobically grown cells, independent of the absence or presence of q. In response to q the LDH A, isoenzyme was further activated by a post-translational mechanism. In q-deficient HeLa cells the activity of the major anoxic stress protein, LDHk, increased as a result of hypoxia; this increase was suppressed by q. In aerobically grown, q-deficient cells significant activities of LDH A, and LDH, were present ; both activities were markedly lowered by q, while the mitochondrial electron flow was improved. The results show that free q is essential for relieving hypoxic stress in HeLa cells that results from oxygen limitation. The modified base queuine [q; 7-5-( [(lS,4S,5R)-4,5-di-anine], is synthesized in eubacteria only. During the biosynthesis GTP is converted to the precursor 7-aminomethyl-7-deazaguanine by so far unknown steps. The tRNA guanine transglycosylase (Tgt) exchanges this precursor specifically for the guanine residue in the anticodons GUN of tRNAs specifying the amino acids Asn, Asp, His and Tyr [l]. The cyclopentendiol moiety is derived from the ribose residue of S-adenosylmethionine and is attached to the modified tRNA precursor by a novel AdoMet-dependent reaction that is catalyzed by the tRNA ribosyltransferase-isomerase (QueA) [2]. During the QueA reaction, epoxy-queuosine is formed that is finally reduced to queuosine (Q) by a vitamin-B ,,-dependent, unidentified enzyme [3]. The genes, tgt and queA, are localized on the Escherichia coli genome in one operon [4].Mammals obtain q from nutrients or the intestinal flora [5-91. Specific eukaryotic Tgt enzymes insert the q-factor in a practically irreversible reaction into the wobble position of cytosolic and of mitochondrial tRNA,,, species [5-71.

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Section: Discussionmentioning

confidence: 99%

The nutrient factor queuine protects HeLa cells from hypoxic stress and improves metabolic adaptation to oxygen availability

Reisser

Langgut

Kersten

1994

European Journal of Biochemistry

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“…Since Q is a nutrient factor for eucaryotes, they contain Q as the free base as well, whereas eubacterial cells do not contain the free Q base. In eucaryotes, Q might be involved in many cellular processes such as differentiation (10,32), the management of hypoxic stress (25,26), the regulation of protein synthesis (25), the adaptive regulation of cell proliferation to the metabolic predisposition (18), and the modulation of signal transduction pathways (17).…”

Section: The Hypermodified Trna Nucleoside Queuosine [Q: 7-(((45-cismentioning

confidence: 99%

Sequence analysis and overexpression of the Zymomonas mobilis tgt gene encoding tRNA-guanine transglycosylase: purification and biochemical characterization of the enzyme

Reuter

Ficner

1995

J Bacteriol

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. We showed that this gene is able to complement the tgt mutation in E. coli SJ1505, and we determined its complete sequence. Four start codons were possible for this gene, resulting in proteins of 386 to 399 amino acids (M r , 42,800 to 44,300) showing 60.4% sequence identity with Tgt from E. coli. The smallest of the four possible reading frames, which was still extended at its 5 end compared with the E. coli tgt gene, was overexpressed in E. coli. The gene product was purified to homogeneity and was biochemically characterized. The kinetical parameters were virtually identical to those published for the E. coli enzyme. In contrast to E. coli Tgt, which is reported to be a homotrimer, Z. mobilis Tgt was found to be a monomer according to gel filtration. In this study, it was shown that the formation of homotrimers by the E. coli enzyme is readily reversible and is dependent on protein concentration. Asn in most eucaryotes and eubacteria. Since Q is a nutrient factor for eucaryotes, they contain Q as the free base as well, whereas eubacterial cells do not contain the free Q base. In eucaryotes, Q might be involved in many cellular processes such as differentiation (10, 32), the management of hypoxic stress (25, 26), the regulation of protein synthesis (25), the adaptive regulation of cell proliferation to the metabolic predisposition (18), and the modulation of signal transduction pathways (17). The hypermodified tRNA nucleoside queuosine [Q: 7-(((4,5-cis-dihydroxy-2-cyclopentene-1-yl)amino)methyl)-7-deaza-In eubacteria, the tRNAs are fully modified with respect to Q under all growth conditions. The exact function of Q in tRNA has not yet been clarified. Escherichia coli mutants deficient in Q biosynthesis die earlier during stationary phase in coculture with wild-type cells (22). They show a significantly higher rate of protein synthesis during outgrowth after nutritional starvation than wild-type cells (36). Thus, the Q in tRNAs seems to participate in the fine tuning of protein biosynthesis, as it is found for other tRNA modifications (2). There is recent evidence for a distinct function of Q in eubacterial tRNAs. A mutation giving rise to apathogenicity in the dysentery-causing bacterium Shigella flexneri was identified as a mutation in a Q biosynthesis gene. In this organism, Q deficiency leads to an inefficient translation of the virF mRNA, which contains the information for a positive regulator of further virulence genes. The mechanism of this phenomenon has not yet been clarified (9).Eubacteria synthesize Q de novo. Biosynthesis (see Fig. 1) starts outside the tRNA, with GTP being converted to 7-aminomethyl-7-deazaguanine (preQ 1 ) by one or several reactions that have not yet been characterized (15). preQ 1 is then inserted into the first position of the anticodon of the tRNA, replacing the genetically encoded guanine. The reaction is catalyzed by the enzyme tRNA-guanine transglycosylase (Tgt [EC 2.4.2.29]) (23). Further biosynthesis occurs at the tRNA level, where a substituted cyclopentyl group deriv...

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“…In contrast to eubacteria, eukaryotes cannot synthesize queuine de novo and are required to obtain queuine from diet or gut flora as a nutrient factor (Reyniers et al 1981). Although its physiological role is largely unclear, queuine modification of tRNA has been suggested to be involved in cell differentiation, proliferation, and response to oxidative stress (Langgut et al 1993;Reisser et al 1994;Pathak et al 2007). In addition, it was reported that queuine-and tyrosine-deficient mice showed severe abnormalities such as labored breathing, seizures, and even death in some cases (Marks and Farkas 1997).…”

Section: Introductionmentioning

confidence: 99%

Characterization of the human tRNA-guanine transglycosylase: Confirmation of the heterodimeric subunit structure

Chen

Kelly

Stachura

et al. 2010

RNA

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The eukaryotic tRNA-guanine transglycosylase (TGT) has been reported to exist as a heterodimer, in contrast to the homodimeric eubacterial TGT. While ubiquitin-specific protease 14 (USP14) has been proposed to act as a regulatory subunit of the eukaryotic TGT, the mouse TGT has recently been shown to be a queuine tRNA-ribosyltransferase 1 (QTRT1, eubacterial TGT homolog) d queuine tRNA-ribosyltransferase domain-containing 1 (QTRTD1) heterodimer. We find that human QTRTD1 (hQTRTD1) co-purifies with polyhistidine-tagged human QTRT1 (ht-hQTRT1) via Ni 2+ affinity chromatography. Cross-linking experiments, mass spectrometry, and size exclusion chromatography results are consistent with the two proteins existing as a heterodimer. We have not been able to observe co-purification and/or association between hQTRT1 and USP14 when coexpressed in Escherichia coli. More importantly, under our experimental conditions, the transglycosylase activity of hQTRT1 is only observed when hQTRT1 and hQTRTD1 have been co-expressed and co-purified. Kinetic characterization of the human TGT (hQTRT1 d hQTRTD1) using human tRNA Tyr and guanine shows catalytic efficiency (k cat /K M ) similar to that of the E. coli TGT. Furthermore, site-directed mutagenesis confirms that the hQTRT1 subunit is responsible for the transglycosylase activity. Taken together, these results indicate that the human TGT is composed of a catalytic subunit, hQTRT1, and hQTRTD1, not USP14. hQTRTD1 has been implicated as the salvage enzyme that generates free queuine from QMP. Work is ongoing in our laboratory to confirm this activity.

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Modulation of mammalian cell proliferation by a modified tRNA base of bacterial origin

Cited by 24 publications

References 24 publications

The nutrient factor queuine protects HeLa cells from hypoxic stress and improves metabolic adaptation to oxygen availability

The nutrient factor queuine protects HeLa cells from hypoxic stress and improves metabolic adaptation to oxygen availability

Sequence analysis and overexpression of the Zymomonas mobilis tgt gene encoding tRNA-guanine transglycosylase: purification and biochemical characterization of the enzyme

Characterization of the human tRNA-guanine transglycosylase: Confirmation of the heterodimeric subunit structure

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