nucleoside-q and queuine

Micronutrients from the diet and gut microbiota are essential to human health and wellbeing. Arguably, among the most intriguing and enigmatic of these micronutrients is queuine, an elaborate 7-deazaguanine derivative made exclusively by eubacteria and salvaged by animal, plant and fungal species. In eubacteria and eukaryotes, queuine is found as the sugar nucleotide queuosine within the anticodon loop of transfer RNA isoacceptors for the amino acids tyrosine, asparagine, aspartic acid and histidine. The physiological requirement for the ancient queuine molecule and queuosine modified transfer RNA has been the subject of varied scientific interrogations for over four decades, establishing relationships to development, proliferation, metabolism, cancer, and tyrosine biosynthesis in eukaryotes and to invasion and proliferation in pathogenic bacteria, in addition to ribosomal frameshifting in viruses. These varied effects may be rationalized by an important, if ill-defined, contribution to protein translation or may manifest from other presently unidentified mechanisms. This article will examine the current understanding of queuine uptake, tRNA incorporation and salvage by eukaryotic organisms and consider some of the physiological consequence arising from deficiency in this elusive and lesser-recognized micronutrient.

Topics: Aging; Animals; Disease Models, Animal; Gastrointestinal Microbiome; Gastrointestinal Tract; Guanine; Humans; Micronutrients; Mitochondria; Neoplasms; Nucleoside Q; RNA, Transfer; Translations

Eukaryotic life benefits from-and ofttimes critically relies upon-the de novo biosynthesis and supply of vitamins and micronutrients from bacteria. The micronutrient queuosine (Q), derived from diet and/or the gut microbiome, is used as a source of the nucleobase queuine, which once incorporated into the anticodon of tRNA contributes to translational efficiency and accuracy. Here, we report high-resolution, substrate-bound crystal structures of the Sphaerobacter thermophilus queuine salvage protein Qng1 (formerly DUF2419) and of its human ortholog QNG1 (C9orf64), which together with biochemical and genetic evidence demonstrate its function as the hydrolase releasing queuine from queuosine-5'-monophosphate as the biological substrate. We also show that QNG1 is highly expressed in the liver, with implications for Q salvage and recycling. The essential role of this family of hydrolases in supplying queuine in eukaryotes places it at the nexus of numerous (patho)physiological processes associated with queuine deficiency, including altered metabolism, proliferation, differentiation and cancer progression.

Topics: Chloroflexi; Glycoside Hydrolases; Guanine; Humans; Micronutrients; Nucleoside Q; Proteins; RNA, Transfer

Transfer RNA (tRNA) is the most extensively modified RNA in cells. Queuosine modification is a fundamental process for ensuring the fidelity and efficiency of translation from RNA to protein. In eukaryotes, Queuosine tRNA (Q-tRNA) modification relies on the intestinal microbial product queuine. However, the roles and potential mechanisms of Q-containing tRNA (Q-tRNA) modifications in inflammatory bowel disease (IBD) are unknown.. We explored the Q-tRNA modifications and expression of QTRT1 (queuine tRNA-ribosyltransferase 1) in patients with IBD by investigating human biopsies and reanalyzing datasets. We used colitis models, QTRT1 knockout mice, organoids, and cultured cells to investigate the molecular mechanisms of Q-tRNA modifications in intestinal inflammation.. QTRT1 expression was significantly downregulated in ulcerative colitis and Crohn's disease patients. The 4 Q-tRNA-related tRNA synthetases (asparaginyl-, aspartyl-, histidyl-, and tyrosyl-tRNA synthetase) were decreased in IBD patients. This reduction was further confirmed in a dextran sulfate sodium-induced colitis model and interleukin-10-deficient mice. Reduced QTRT1 was significantly correlated with cell proliferation and intestinal junctions, including downregulation of β-catenin and claudin-5 and the upregulation of claudin-2. These alterations were confirmed in vitro by deleting the QTRT1 gene from cells and in vivo using QTRT1 knockout mice. Queuine treatment significantly enhanced cell proliferation and junction activity in cell lines and organoids. Queuine treatment also reduced inflammation in epithelial cells. Moreover, altered QTRT1-related metabolites were found in human IBD.. tRNA modifications play an unexplored novel role in the pathogenesis of intestinal inflammation by altering epithelial proliferation and junction formation. Further investigation of the role of tRNA modifications will uncover novel molecular mechanisms for the prevention and treatment of IBD.

Topics: Animals; Colitis; Humans; Inflammation; Inflammatory Bowel Diseases; Mice; Mice, Knockout; Nucleoside Q; RNA, Transfer

Queuosine (Q) is a hypermodified 7-deaza-guanosine nucleoside that is found at position 34, also known as the wobble position, of tRNAs with a GUN anticodon, and Q ensures faithful translation of the respective C- and U-ending codons. While Q is present in tRNAs in most eukaryotes, only bacteria can synthesize it denovo. In contrast, eukaryotes rely on external sources like their food and the gut microbiome in order to Q-modify their tRNAs, and Q therefore can be regarded as a micronutrient. The eukaryotic tRNA guanine transglycosylase (eTGT) uses the base queuine (q) as a substrate to replace G34 by Q in the tRNAs. Eukaryotic cells can uptake both q and Q, raising the question how the Q nucleoside is converted to q for incorporation into the tRNAs. Here, we identified Qng1 (also termed Duf2419) as a queuosine nucleoside glycosylase in Schizosaccharomyces pombe. S. pombe cells with a deletion of qng1

Topics: Animals; Bacteria; Guanine; Humans; Hydrolysis; Mice; Nucleoside Q; Nucleosides; RNA, Transfer; Schizosaccharomyces

Every type of nucleic acid in cells undergoes programmed chemical post-transcriptional modification. Generally, modification enzymes use substrates derived from intracellular metabolism, one exception is queuine (q)/queuosine (Q), which eukaryotes obtain from their environment; made by bacteria and ultimately taken into eukaryotic cells via currently unknown transport systems. Here, we use a combination of molecular, cell biology and biophysical approaches to show that in Trypanosoma brucei tRNA Q levels change dynamically in response to concentration variations of a sub-set of amino acids in the growth media. Most significant were variations in tyrosine, which at low levels lead to increased Q content for all the natural tRNAs substrates of tRNA-guanine transglycosylase (TGT). Such increase results from longer nuclear dwell time aided by retrograde transport following cytoplasmic splicing. In turn high tyrosine levels lead to rapid decrease in Q content. Importantly, the dynamic changes in Q content of tRNAs have negligible effects on global translation or growth rate but, at least, in the case of tRNATyr it affected codon choice. These observations have implications for the occurrence of other tunable modifications important for 'normal' growth, while connecting the intracellular localization of modification enzymes, metabolites and tRNAs to codon selection and implicitly translational output.

Topics: Amino Acids; Chromatography, Liquid; Codon; Guanine; Nucleoside Q; Nutrients; Pentosyltransferases; Protozoan Proteins; RNA Splicing; RNA, Transfer; RNA, Transfer, Tyr; Tandem Mass Spectrometry; Trypanosoma brucei brucei; Tyrosine

Queuosine-deficient tRNAs are often observed in neoplastic cells. In order to determine possible sites for malfunction of the multistep queuosine modification system, comprehensive studies were performed on two human neoplastic cell lines, the HxGC(3) colon adenocarcinoma and the MCF-7 breast adenocarcinoma, which are 100 and 50-60% queuosine deficient, respectively. These results were compared with data obtained from normal human fibroblast (HFF) cultures which maintain 100% queuosine-modified tRNA populations. Queuine uptake in all three cell types was similar and each demonstrated activation by protein kinase C (PKC). However, incorporation of queuine into tRNA by tRNA:guanine ribosyltransferase (TGRase; E.C. 2.4.2.24) and PKC-catalyzed activation of this enzyme occurred only in HFF and MCF-7 cells. The HxGC(3) cell line exhibited no TGRase activity as was expected. Treatment with 5-azacytidine (5-azaC) induced TGRase activity to a level 20% of that in HFF and MCF-7 cells; however, this 5-azaC-induced TGRase activity was not regulated by PKC. Salvage of the queuine base from tRNA degradation products has been shown in mammalian cells and was measured in the HFF cells. However, salvage activity in the MCF-7 cell line was deficient. Therefore, it was shown by direct measurements that the HxGC(3) cell line is completely lacking in queuosine-modified tRNA due to loss of functional TGRase, while the MCF-7 cell line has an inefficient queuine salvage mechanism resulting in a significant deficiency of queuosine-modified tRNA. These techniques can be applied to any cultured cell types to determine specific lesions of the queuosine modification system, which have been suggested to be associated with neoplastic progression.

Topics: Azacitidine; Cells, Cultured; Enzyme Inhibitors; Guanine; Humans; Male; Nucleoside Q; Phosphorylation; Protein Kinase C; RNA, Transfer; Staurosporine; Tetradecanoylphorbol Acetate; Time Factors; Tumor Cells, Cultured

Queuosine (Q) [7-(((4,5-cis-dihydroxy-2-cyclopenten-1-yl)amino)methyl)-7-deaz agu anosine] usually occurs in the first position of the anticodon of tRNAs specifying the amino acids asparagine, aspartate, histidine, and tyrosine. The hypermodified nucleoside is found in eubacteria and eucaryotes. Q is synthesized de novo exclusively in eubacteria; for eucaryotes the compound is a nutrient factor. In Escherichia coli the Q precursor (oQ), carrying a 2,3-epoxy-4,5-dihydroxycyclopentane ring, is formed from tRNA precursors containing 7-(aminomethyl)-7-deazaguanine (preQ1) by the queA gene product. A genomic queA mutant accumulating preQ1 tRNA was constructed. The QueA enzyme was overexpressed as a fusion protein with the glutathione S-transferase from Schistosoma japonicum and purified to homogeneity by affinity and anion-exchange chromatography. The enzyme QueA synthesizes oQ from preQ1 in a single S-adenosylmethionine- (AdoMet-) requiring step, indicating that the ribosyl moiety of AdoMet is transferred and isomerized to the epoxycyclopentane residue of oQ. The identity of oQ was verified by HPLC and directly combined HPLC/mass spectrometry. The formation of oQ was reconstituted in vitro, applying a synthetic RNA. A 17-nucleotide microhelix (corresponding to the anticodon stem and loop of tRNA(Tyr) from E. coli) is sufficient to act as the RNA substrate for oQ synthesis. We propose that QueA is an S-adenosylmethionine:tRNA ribosyltransferase-isomerase.

Topics: Amino Acid Sequence; Animals; Base Sequence; Chromatography, Gel; Chromatography, High Pressure Liquid; Electrophoresis, Polyacrylamide Gel; Enzyme Precursors; Escherichia coli; Guanine; Isomerases; Molecular Sequence Data; Nucleoside Q; Pentosyltransferases; RNA, Transfer, Tyr; S-Adenosylmethionine; Schistosoma japonicum

Partially purified extracts from Chlorella pyrenoidosa and Chlamydomonas reinhardtii catalyze the cleavage of queuosine (Q), a modified 7-deazaguanine nucleoside found exclusively in the first position of the anticodon of certain tRNAs, to queuine, the base of Q. This is the first report of an enzyme that specifically cleaves a 7-deazapurine riboside. Guanosine is not a substrate for this activity, nor is the epoxide a derivative of Q. We also establish that both algae can incorporate exogenously supplied queuine into their tRNA but lack Q-containing tRNA when cultivated in the absence of queuine, indicating that they are unable to synthesize Q de novo. Although no physiological function for Q has been identified in these algae, Q cleavage to queuine would enable algae to generate queuine from exogenous Q in the wild and also to salvage (and recycle) queuine from intracellular tRNA degraded during the normal turnover process. In mammalian cells, queuine salvage occurs by the specific cleavage of queuine from Q-5'-phosphate. The present data also support the hypothesis that plants, like animals, cannot synthesize Q de novo.

Topics: Chlamydomonas; Chlorella; Chromatography, High Pressure Liquid; Chromatography, Liquid; Guanine; Guanosine; Mass Spectrometry; Nucleoside Q; RNA, Transfer; Species Specificity

Topics: Animals; Cell Line; Choriocarcinoma; Diet; Female; Guanine; Guanosine; Kidney; Male; Mice; Neoplasms; Neuroblastoma; Nucleoside Q; Pregnancy; RNA, Transfer; Species Specificity

1. Normal mouse liver contains predominantly tRNA that contains queuosine in the first position of the anticodon ((Q+)tRNA). 2. Germ-free mice fed a chemically defined diet devoid of queuine for 1 year have no queuine in all four of the tRNAs that respond to the NAUC codons. 3. The synthesis of (Q+)tRNAs can be induced by injecting queuine, feeding free queuine, or by feeding (Q+)tRNA. 4. When mice that have no (Q+)tRNA are titrated with exogenous queuine, tRNAAsp is modified to the (Q+) state before tRNAHis.

Topics: Animals; Guanine; Guanosine; Liver; Mice; Mice, Inbred Strains; Nucleoside Q; RNA, Transfer

Topics: Amniotic Fluid; Animals; Carcinoma, Ehrlich Tumor; Cattle; Cell Division; Female; Guanine; Guanosine; Histidine; Mice; Nucleoside Q; Pregnancy; RNA, Transfer