Queuosine Deficiency in Eukaryotes Compromises Tyrosine Production through Increased Tetrahydrobiopterin Oxidation

Abstract

Queuosine is a modified pyrrolopyrimidine nucleoside found in the anticodon loop of transfer RNA acceptors for the amino acids tyrosine, asparagine, aspartic acid, and histidine. Because it is exclusively synthesized by bacteria, higher eukaryotes must salvage queuosine or its nucleobase queuine from food and the gut microflora. Previously, animals made deficient in queuine died within 18 days of withdrawing tyrosine, a nonessential amino acid, from the diet (Marks, T., and Farkas, W. R. (1997) Biochem. Biophys. Res. Commun. 230, 233-237). Here, we show that human HepG2 cells deficient in queuine and mice made deficient in queuosine-modified transfer RNA, by disruption of the tRNA guanine transglycosylase enzyme, are compromised in their ability to produce tyrosine from phenylalanine. This has similarities to the disease phenylketonuria, which arises from mutation in the enzyme phenylalanine hydroxylase or from a decrease in the supply of its cofactor tetrahydrobiopterin (BH4). Immunoblot and kinetic analysis of liver from tRNA guanine transglycosylase-deficient animals indicates normal expression and activity of phenylalanine hydroxylase. By contrast, BH4 levels are significantly decreased in the plasma, and both plasma and urine show a clear elevation in dihydrobiopterin, an oxidation product of BH4, despite normal activity of the salvage enzyme dihydrofolate reductase. Our data suggest that queuosine modification limits BH4 oxidation in vivo and thereby potentially impacts on numerous physiological processes in eukaryotes.Bacteria and humans have co-evolved for millennia, and many examples exist of how various symbiotic and commensal partnerships contribute to human health and nutrition ranging from the metabolism of complex carbohydrates to the provision of vital micronutrients (1). Queuosine is an example of a micronutrient, synthesized exclusively by bacteria but which, for poorly defined reasons, is utilized by almost all eukaryotic species with the exception of the baker's yeast, Saccharomyces cerevisiae (2).Bacterial queuosine biosynthesis occurs in two stages. First, a series of five enzymatic steps convert guanosine triphosphate nucleoside (GTP) to the soluble 7-aminomethyl-7-deazaguanine molecule. Subsequently, 7-aminomethyl-7-deazaguanine is inserted into the wobble position of tRNA containing a GUN consensus sequence (Tyr, Asp, Asn, and His) by means of the single enzyme species, tRNA guanine transglycosylase (TGT), and is further remodeled in situ to queuosine (3). Eukaryotes must acquire queuosine or its free nucleobase, queuine, from food and the gut microflora. Curiously, both cytosolic and mitochondrial tRNA species are modified by queuosine (2). The eukaryotic enzyme that performs this reaction, queuine tRNA ribosyltransferase, has recently been identified as a heterodimeric complex, consisting of the eukaryotic homologue of the catalytic TGT subunit and a related protein called queuine tRNA ribosyltransferase domain containing 1 (QTRTD1), both of which localize to the mitochondria (4, 5).Stu...

Structural, Functional, and Taxonomic Diversity of Three PreQ1 Riboswitch Classes

“…PreQ 1 and Q biosynthesis genes are required for effective nitrogen-fixing symbiosis between Sinorhizobium meliloti and the barrel clover plant (Marchetti et al, 2013). Also, glutamyl-Q, another Q derivative, has been linked to effective stress response in S. flexneri (Caballero et al, 2012), while Q-deficiency halts tyrosine production in germ-free mice (Rakovich et al, 2011). In human and murine cancer cells, lack of Q-modification in tRNAs has been associated with cell proliferation, anaerobic metabolism, and malignancy (Elliot et al, 1984; Pathak et al, 2008).…”

“…PreQ 1 is a guanine-derived nucleobase ( Figure 1A ) that is known to be incorporated in the wobble position of tRNAs containing the GUN anticodon sequence and then further modified to yield queuosine (Q) (Iwata-Reuyl, 2008). The presence of Q in tRNAs improves their ability to read degenerate codons (Meier et al, 1985), while the absence of enzymes involved in Q biosynthesis or salvage results in deleterious phenotypes in a wide array of organisms (Durand et al, 2000; Rakovich et al, 2011). Despite the near-ubiquitous presence of preQ 1 in organisms (Iwata-Reuyl, 2008), much remains unknown regarding the function of this modified nucleobase and its derivatives in cells.…”

Biosynthesis of pyrrolopyrimidines

“…A recent study demonstrated that liver extracts from mice that lack the enzymes for incorporating Q into RNA contained decreased levels of tetrahydrobiopterin, the cofactor required for the conversion of phenylalanine to tyrosine by phenylalanine hydroxylase [31]. …”

Major reorientation of tRNA substrates defines specificity of dihydrouridine synthases

“…Altered levels of tRNA modification have been linked to several disorders including cancers (2)(3)(4)(5)(6)(7)(8). One of the most common modified nucleosides, dihydrouridine, is produced by reduction of the C5-C6 double bond in uridine.…”