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Transfer RNA (tRNA) modifications have emerged as critical posttranscriptional regulators of gene expression affecting diverse biological and disease processes. While there is extensive knowledge about the enzymes installing the dozens of post-transcriptional tRNA modifications - the tRNA epitranscriptome - very little is known about how metabolic, signaling, and other networks integrate to regulate tRNA modification levels. Here we took a comprehensive first step at understanding epitranscriptome regulatory networks by developing a high-throughput tRNA isolation and mass spectrometry-based modification profiling platform and applying it to a Pseudomonas aeruginosa transposon insertion mutant library comprising 5,746 strains. Analysis of >200,000 tRNA modification data points validated the annotations of predicted tRNA modification genes, uncovered novel tRNA-modifying enzymes, and revealed tRNA modification regulatory networks in P. aeruginosa. Platform adaptation for RNA-seq library preparation would complement epitranscriptome studies, while application to human cell and mouse tissue demonstrates its utility for biomarker and drug discovery and development.
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Multidrug-resistant P. aeruginosa infections pose a serious public health threat due to the rise in antimicrobial resistance. Phage therapy has emerged as a promising alternative. However, P. aeruginosa has evolved various mechanisms to thwart phage attacks, making it crucial to decipher these resistance mechanisms to develop effective therapeutic strategies. In this study, we conducted a forward-genetic screen of the P. aeruginosa PA14 non-redundant transposon library (PA14NR) to identify dominant-negative mutants displaying phage-resistant phenotypes. Our screening process revealed 78 mutants capable of thriving in the presence of phages, with 23 of them carrying insertions in genes associated with membrane composition. Six mutants exhibited total resistance to phage infection. Transposon insertions were found in genes known to be linked to phage-resistance such as galU and a glycosyl transferase gene, as well as novel genes such as mexB, lasB, and two hypothetical proteins. Functional experiments demonstrated that these genes played pivotal roles in phage adsorption and biofilm formation, indicating that altering the bacterial membrane composition commonly leads to phage resistance in P. aeruginosa. Importantly, these mutants displayed phenotypic trade-offs, as their resistance to phages inversely affected antibiotic resistance and hindered biofilm formation, shedding light on the complex interplay between phage susceptibility and bacterial fitness. This study highlights the potential of transposon mutant libraries and forward-genetic screens in identifying key genes involved in phage-host interactions and resistance mechanisms. These findings support the development of innovative strategies for combating antibiotic-resistant pathogens.
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Elementos de DNA Transponíveis , Biblioteca Gênica , Mutação , Pseudomonas aeruginosa , Pseudomonas aeruginosa/virologia , Pseudomonas aeruginosa/genética , Elementos de DNA Transponíveis/genética , Biofilmes/crescimento & desenvolvimento , Bacteriófagos/genética , Bacteriófagos/fisiologiaRESUMO
Mycobacteria adapt to infection stresses by entering a reversible non-replicating persistence (NRP) with slow or no cell growth and broad antimicrobial tolerance. Hypoxia and nutrient deprivation are two well-studied stresses commonly used to model the NRP, yet little is known about the molecular differences in mycobacterial adaptation to these distinct stresses that lead to a comparable NRP phenotype. Here we performed a multisystem interrogation of the Mycobacterium bovis BCG (BCG) starvation response, which revealed a coordinated metabolic shift away from the glycolysis of nutrient-replete growth to depletion of lipid stores, lipolysis, and fatty acid ß-oxidation in NRP. This contrasts with BCG's NRP hypoxia response involving a shift to cholesterol metabolism and triglyceride storage. Our analysis reveals cryptic metabolic vulnerabilities of the starvation-induced NRP state, such as their newfound hypersensitivity to H2O2. These observations pave the way for developing precision therapeutics against these otherwise drug refractory pathogens.
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Adaptação Fisiológica , Mycobacterium bovis , Mycobacterium bovis/metabolismo , Glicólise , Reprogramação MetabólicaRESUMO
Among dozens of known epigenetic marks, naturally occurring phosphorothioate (PT) DNA modifications are unique in replacing a non-bridging phosphate oxygen with redox-active sulfur and function in prokaryotic restriction-modification and transcriptional regulation. Interest in PTs has grown due to the widespread distribution of the dnd, ssp, and brx genes among bacteria and archaea, as well as the discovery of PTs in 5-10% of gut microbes. Efforts to map PTs in complex microbiomes using existing next-generation and direct sequencing technologies have failed due to poor sensitivity. Here we developed PT-seq as a high-sensitivity method to quantitatively map PTs across genomes and metagenomically identify PT-containing microbes in complex genomic mixtures. Like other methods for mapping PTs in individual genomes, PT-seq exploits targeted DNA strand cleavage at PTs by iodine, followed by sequencing library construction using ligation or template switching approaches. However, PT-specific sequencing reads are dramatically increased by adding steps to heat denature the DNA, block pre-existing 3'-ends, fragment DNA after T-tailing, and enrich iodine-induced breaks using biotin-labeling and streptavidin beads capture. Iterative optimization of the sensitivity and specificity of PT-seq is demonstrated with individual bacteria and human fecal DNA.
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Background: Epigenetic regulation of gene expression and host defense is well established in microbial communities, with dozens of DNA modifications comprising the epigenomes of prokaryotes and bacteriophage. Phosphorothioation (PT) of DNA, in which a chemically-reactive sulfur atom replaces a non-bridging oxygen in the sugar-phosphate backbone, is catalyzed by dnd and ssp gene families widespread in bacteria and archaea. However, little is known about the role of PTs or other microbial epigenetic modifications in the human microbiome. Here we optimized and applied fecal DNA extraction, mass spectrometric, and metagenomics technologies to characterize the landscape and temporal dynamics of gut microbes possessing PT modifications. Results: Exploiting the nuclease-resistance of PTs, mass spectrometric analysis of limit digests of PT-containing DNA reveals PT dinucleotides as part of genomic consensus sequences, with 16 possible dinucleotide combinations. Analysis of mouse fecal DNA revealed a highly uniform spectrum of 11 PT dinucleotides in all littermates, with PTs estimated to occur in 5-10% of gut microbes. Though at similar levels, PT dinucleotides in fecal DNA from 11 healthy humans possessed signature combinations and levels of individual PTs. Comparison with a widely distributed microbial epigenetic mark, m6dA, suggested temporal dynamics consistent with expectations for gut microbial communities based on Taylor's Power Law. Application of PT-seq for site-specific metagenomic analysis of PT-containing bacteria in one fecal donor revealed the larger consensus sequences for the PT dinucleotides in Bacteroidota, Firmicutes, Actinobacteria, and Proteobacteria, which differed from unbiased metagenomics and suggested that the abundance of PT-containing bacteria did not simply mirror the spectrum of gut bacteria. PT-seq further revealed low abundance PT sites not detected as dinucleotides by mass spectrometry, attesting to the complementarity of the technologies. Conclusions: The results of our studies provide a benchmark for understanding the behavior of an abundant and chemically-reactive epigenetic mark in the human gut microbiome, with implications for inflammatory conditions of the gut.
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Among dozens of microbial DNA modifications regulating gene expression and host defense, phosphorothioation (PT) is the only known backbone modification, with sulfur inserted at a non-bridging oxygen by dnd and ssp gene families. Here we explored the distribution of PT genes in 13,663 human gut microbiome genomes, finding that 6.3% possessed dnd or ssp genes predominantly in Bacillota, Bacteroidota, and Pseudomonadota. This analysis uncovered several putative new PT synthesis systems, including Type 4 Bacteriophage Exclusion (BREX) brx genes, which were genetically validated in Bacteroides salyersiae. Mass spectrometric analysis of DNA from 226 gut microbiome isolates possessing dnd, ssp, and brx genes revealed 8 PT dinucleotide settings confirmed in 6 consensus sequences by PT-specific DNA sequencing. Genomic analysis showed PT enrichment in rRNA genes and depletion at gene boundaries. These results illustrate the power of the microbiome for discovering prokaryotic epigenetics and the widespread distribution of oxidation-sensitive PTs in gut microbes.
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Tens of thousands of people in southern Europe suffer from Balkan endemic nephropathy (BEN), and four times as many are at risk. Incidental ingestion of aristolochic acids (AAs), stemming from the ubiquitousAristolochia clematitis(birthwort) weed in the region, leads to DNA adduct-induced toxicity in kidney cells, the primary cause of BEN. Numerous cofactors, including toxic organics and metals, have been investigated, but all have shown small contributions to the overall BEN relative to non-BEN village distribution gradients. Here, we reveal that combustion-derived pollutants from wood and coal burning in Serbia also contaminate arable soil and test as plausible causative factors of BEN. Using a GC-MS screening method, biomass-burning-derived furfural and coal-burning-derived medium-chain alkanes were detected in soil samples from BEN endemic areas levels at up to 63-times and 14-times higher, respectively, than in nonendemic areas. Significantly higher amounts were also detected in colocated wheat grains. Coexposure studies with cultured kidney cells showed that these pollutants enhance DNA adduct formation by AA, - the cause of AA nephrotoxicity and carcinogenicity. With the coincidence of birthwort-derived AAs and the widespread practice of biomass and coal burning for household cooking and heating purposes and agricultural burning in rural low-lying flood-affected areas in the Balkans, these results implicate combustion-derived pollutants in promoting the development of BEN.
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Nefropatia dos Bálcãs , Inundações , Nefropatia dos Bálcãs/induzido quimicamente , Nefropatia dos Bálcãs/epidemiologia , Humanos , Carvão Mineral , Sérvia , Poluentes do Solo/toxicidade , Ácidos Aristolóquicos , Animais , Aristolochia/química , Península Balcânica , Madeira , Nefropatias/induzido quimicamenteRESUMO
Ribonucleoside modifications comprising the epitranscriptome are present in all organisms and all forms of RNA, including mRNA, rRNA and tRNA, the three major RNA components of the translational machinery. Of these, tRNA is the most heavily modified and the tRNA epitranscriptome has the greatest diversity of modifications. In addition to their roles in tRNA biogenesis, quality control, structure, cleavage, and codon recognition, tRNA modifications have been shown to regulate gene expression post-transcriptionally in prokaryotes and eukaryotes, including humans. However, studies investigating the impact of tRNA modifications on gene expression in the malaria parasite Plasmodium falciparum are currently scarce. Current evidence shows that the parasite has a limited capacity for transcriptional control, which points to a heavier reliance on strategies for posttranscriptional regulation such as tRNA epitranscriptome reprogramming. This review addresses the known functions of tRNA modifications in the biology of P. falciparum while highlighting the potential therapeutic opportunities and the value of using P. falciparum as a model organism for addressing several open questions related to the tRNA epitranscriptome.
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Plasmodium falciparum artemisinin (ART) resistance is driven by mutations in kelch-like protein 13 (PfK13). Quiescence, a key aspect of resistance, may also be regulated by a yet unidentified epigenetic pathway. Transfer RNA modification reprogramming and codon bias translation is a conserved epitranscriptomic translational control mechanism that allows cells to rapidly respond to stress. We report a role for this mechanism in ART-resistant parasites by combining tRNA modification, proteomic and codon usage analyses in ring-stage ART-sensitive and ART-resistant parasites in response to drug. Post-drug, ART-resistant parasites differentially hypomodify mcm5s2U on tRNA and possess a subset of proteins, including PfK13, that are regulated by Lys codon-biased translation. Conditional knockdown of the terminal s2U thiouridylase, PfMnmA, in an ART-sensitive parasite background led to increased ART survival, suggesting that hypomodification can alter the parasite ART response. This study describes an epitranscriptomic pathway via tRNA s2U reprogramming that ART-resistant parasites may employ to survive ART-induced stress.
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Antimaláricos , Artemisininas , Resistência a Medicamentos , Plasmodium falciparum , Proteínas de Protozoários , RNA de Transferência , Plasmodium falciparum/genética , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/metabolismo , Resistência a Medicamentos/genética , RNA de Transferência/genética , RNA de Transferência/metabolismo , Artemisininas/farmacologia , Antimaláricos/farmacologia , Proteínas de Protozoários/genética , Proteínas de Protozoários/metabolismo , Humanos , Malária Falciparum/parasitologia , Proteômica , Códon/genéticaRESUMO
Introduction: The study of the pathophysiology study of Alzheimer's disease (AD) has been hampered by lack animal models that recapitulate the major AD pathologies, including extracellular -amyloid (A) deposition, intracellular aggregation of microtubule associated protein tau (MAPT), inflammation and neurodegeneration. Methods: The humanized APPNL-G-F knock-in mouse line was crossed to the PS19 MAPTP301S, over-expression mouse line to create the dual APPNL-G-F/PS19 MAPTP301S line. The resulting pathologies were characterized by immunochemical methods and PCR. Results: We now report on a double transgenic APPNL-G-F/PS19 MAPTP301S mouse that at 6 months of age exhibits robust A plaque accumulation, intense MAPT pathology, strong inflammation and extensive neurodegeneration. The presence of A pathology potentiated the other major pathologies, including MAPT pathology, inflammation and neurodegeneration. MAPT pathology neither changed levels of amyloid precursor protein nor potentiated A accumulation. Interestingly, study of immunofluorescence in cleared brains indicates that microglial inflammation was generally stronger in the hippocampus, dentate gyrus and entorhinal cortex, which are regions with predominant MAPT pathology. The APPNL-G-F/MAPTP301S mouse model also showed strong accumulation of N6-methyladenosine (m6A), which was recently shown to be elevated in the AD brain. m6A primarily accumulated in neuronal soma, but also co-localized with a subset of astrocytes and microglia. The accumulation of m6A corresponded with increases in METTL3 and decreases in ALKBH5, which are enzymes that add or remove m6A from mRNA, respectively. Discussion: Our understanding of the pathophysiology of Alzheimer's disease (AD) has been hampered by lack animal models that recapitulate the major AD pathologies, including extracellular -amyloid (A) deposition, intracellular aggregation of microtubule associated protein tau (MAPT), inflammation and neurodegeneration. The APPNL-G-F/MAPTP301S mouse recapitulates many features of AD pathology beginning at 6 months of aging, and thus represents a useful new mouse model for the field.
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The wobble bases of tRNAs that decode split codons are often heavily modified. In bacteria, tRNAGlu, Gln, Asp contains a variety of xnm5s2U derivatives. The synthesis pathway for these modifications is complex and fully elucidated only in a handful of organisms, including the Gram-negative Escherichia coli K12 model. Despite the ubiquitous presence of mnm5s2U modification, genomic analysis shows the absence of mnmC orthologous genes, suggesting the occurrence of alternate biosynthetic schemes for the conversion of cmnm5s2U to mnm5s2U. Using a combination of comparative genomics and genetic studies, a member of the YtqA subgroup of the radical Sam superfamily was found to be involved in the synthesis of mnm5s2U in both Bacillus subtilis and Streptococcus mutans. This protein, renamed MnmL, is encoded in an operon with the recently discovered MnmM methylase involved in the methylation of the pathway intermediate nm5s2U into mnm5s2U in B. subtilis. Analysis of tRNA modifications of both S. mutans and Streptococcus pneumoniae shows that growth conditions and genetic backgrounds influence the ratios of pathway intermediates owing to regulatory loops that are not yet understood. The MnmLM pathway is widespread along the bacterial tree, with some phyla, such as Bacilli, relying exclusively on these two enzymes. Although mechanistic details of these newly discovered components are not fully resolved, the occurrence of fusion proteins, alternate arrangements of biosynthetic components, and loss of biosynthetic branches provide examples of biosynthetic diversity to retain a conserved tRNA modification in Nature.IMPORTANCEThe xnm5s2U modifications found in several tRNAs at the wobble base position are widespread in bacteria where they have an important role in decoding efficiency and accuracy. This work identifies a novel enzyme (MnmL) that is a member of a subgroup of the very versatile radical SAM superfamily and is involved in the synthesis of mnm5s2U in several Gram-positive bacteria, including human pathogens. This is another novel example of a non-orthologous displacement in the field of tRNA modification synthesis, showing how different solutions evolve to retain U34 tRNA modifications.
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Escherichia coli K12 , RNA de Transferência , Humanos , RNA de Transferência/genética , Escherichia coli K12/genética , Bactérias/genética , Metilação , Bactérias Gram-Positivas/genéticaRESUMO
Transfer RNA (tRNA) modifications play a crucial role in maintaining translational fidelity and efficiency, and they may function as regulatory elements in stress response and virulence. Despite their pivotal roles, a comprehensive mapping of tRNA modifications and their associated synthesis genes is still limited, with a predominant focus on free-living bacteria. In this study, we employed a multidisciplinary approach, incorporating comparative genomics, mass spectrometry, and next-generation sequencing, to predict the set of tRNA modification genes responsible for tRNA maturation in two intracellular pathogens-Bartonella henselae Houston I and Bartonella quintana Toulouse, which are causative agents of cat-scratch disease and trench fever, respectively. This analysis presented challenges, particularly because of host RNA contamination, which served as a potential source of error. However, our approach predicted 26 genes responsible for synthesizing 23 distinct tRNA modifications in B. henselae and 22 genes associated with 23 modifications in B. quintana. Notably, akin to other intracellular and symbiotic bacteria, both Bartonella species have undergone substantial reductions in tRNA modification genes, mostly by simplifying the hypermodifications present at positions 34 and 37. Bartonella quintana exhibited the additional loss of four modifications and these were linked to examples of gene decay, providing snapshots of reductive evolution.
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The role of translational regulation in brown adipogenesis is relatively unknown. Localized translation of mRNAs encoding mitochondrial components enables swift mitochondrial responses, but whether this occurs during brown adipogenesis, which involves massive mitochondrial biogenesis, has not been explored. Here, we used ribosome profiling and RNA-Seq, coupled with cellular fractionation, to obtain spatiotemporal insights into translational regulation. During brown adipogenesis, a translation bias towards G/C-ending codons is triggered first in the mitochondrial vicinity by reactive oxygen species (ROS), which later spreads to the rest of the cell. This translation bias is induced through ROS modulating the activity of the tRNA modification enzyme, ELP3. Intriguingly, functionally relevant mRNAs, including those encoding ROS scavengers, benefit from this bias; in so doing, ROS-induced translation bias both fuels differentiation and concurrently minimizes oxidative damage. These ROS-induced changes could enable sustained mitochondrial biogenesis during brown adipogenesis, and explain in part, the molecular basis for ROS hormesis.
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AQRNA-seq provides a direct linear relationship between sequencing read counts and small RNA copy numbers in a biological sample, thus enabling accurate quantification of the pool of small RNAs. The AQRNA-seq library preparation procedure described here involves the use of custom-designed sequencing linkers and a step for reducing methylation RNA modifications that block reverse transcription processivity, which results in an increased yield of full-length cDNAs. In addition, a detailed implementation of the accompanying bioinformatics pipeline is presented. This demonstration of AQRNA-seq was conducted through a quantitative analysis of the 45 tRNAs in Mycobacterium bovis BCG harvested on 5 selected days across a 20-day time course of nutrient deprivation and 6 days of resuscitation. Ongoing efforts to improve the efficiency and rigor of AQRNA-seq will also be discussed here. This includes exploring methods to obviate gel purification for mitigating primer dimer issues after PCR amplification and to increase the proportion of full-length reads to enable more accurate read mapping. Future enhancements to AQRNA-seq will be focused on facilitating automation and high-throughput implementation of this technology for quantifying all small RNA species in cell and tissue samples from diverse organisms.
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Sequenciamento de Nucleotídeos em Larga Escala , RNA , Sequenciamento de Nucleotídeos em Larga Escala/métodos , RNA/genética , RNA de Transferência/genética , Biblioteca Gênica , DNA Complementar/genética , Análise de Sequência de RNA/métodosRESUMO
Mitochondrial stress and dysfunction play important roles in many pathologies. However, how cells respond to mitochondrial stress is not fully understood. Here, we examined the translational response to electron transport chain (ETC) inhibition and arsenite induced mitochondrial stresses. Our analysis revealed that during mitochondrial stress, tRNA modifications (namely f5C, hm5C, queuosine and its derivatives, and mcm5U) dynamically change to fine tune codon decoding, usage, and optimality. These changes in codon optimality drive the translation of many pathways and gene sets, such as the ATF4 pathway and selenoproteins, involved in the cellular response to mitochondrial stress. We further examined several of these modifications using targeted approaches. ALKBH1 knockout (KO) abrogated f5C and hm5C levels and led to mitochondrial dysfunction, reduced proliferation, and impacted mRNA translation rates. Our analysis revealed that tRNA queuosine (tRNA-Q) is a master regulator of the mitochondrial stress response. KO of QTRT1 or QTRT2, the enzymes responsible for tRNA-Q synthesis, led to mitochondrial dysfunction, translational dysregulation, and metabolic alterations in mitochondria-related pathways, without altering cellular proliferation. In addition, our analysis revealed that tRNA-Q loss led to a domino effect on various tRNA modifications. Some of these changes could be explained by metabolic profiling. Our analysis also revealed that utilizing serum deprivation or alteration with Queuine supplementation to study tRNA-Q or stress response can introduce various confounding factors by altering many other tRNA modifications. In summary, our data show that tRNA modifications are master regulators of the mitochondrial stress response by driving changes in codon decoding.
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Transfer RNA (tRNA) modifications play a crucial role in maintaining translational fidelity and efficiency, and they may function as regulatory elements in stress response and virulence. Despite their pivotal roles, a comprehensive mapping of tRNA modifications and their associated synthesis genes is still limited, with a predominant focus on free-living bacteria. In this study, we employed a multidisciplinary approach, incorporating comparative genomics, mass spectrometry, and next-generation sequencing, to predict the set of tRNA modification genes responsible for tRNA maturation in two intracellular pathogens- Bartonella henselae Houston I and Bartonella quintana Toulouse, which are causative agents of cat-scratch disease and trench fever, respectively. This analysis presented challenges, particularly because of host RNA contamination, which served as a potential source of error. However, our approach predicted 26 genes responsible for synthesizing 23 distinct tRNA modifications in B. henselae and 22 genes associated with 23 modifications in B. quintana . Notably, akin to other intracellular and symbiotic bacteria, both Bartonella species have undergone substantial reductions in tRNA modification genes, mostly by simplifying the hypermodifications present at positions 34 and 37. B. quintana exhibited the additional loss of four modifications and these were linked to examples of gene decay, providing snapshots of reductive evolution.
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Antibodies are produced at high rates to provide immunoprotection, which puts pressure on the B cell translational machinery. Here, we identified a pattern of codon usage conserved across antibody genes. One feature thereof is the hyperutilization of codons that lack genome-encoded Watson-Crick transfer RNAs (tRNAs), instead relying on the posttranscriptional tRNA modification inosine (I34), which expands the decoding capacity of specific tRNAs through wobbling. Antibody-secreting cells had increased I34 levels and were more reliant on I34 for protein production than naïve B cells. Furthermore, antibody I34-dependent codon usage may influence B cell passage through regulatory checkpoints. Our work elucidates the interface between the tRNA pool and protein production in the immune system and has implications for the design and selection of antibodies for vaccines and therapeutics.
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Anticorpos , Formação de Anticorpos , Linfócitos B , Uso do Códon , Cadeias Pesadas de Imunoglobulinas , Inosina , RNA de Transferência , Formação de Anticorpos/genética , Códon/genética , Inosina/genética , Inosina/metabolismo , RNA de Transferência/genética , Anticorpos/genética , Humanos , Linfócitos B/imunologia , Cadeias Pesadas de Imunoglobulinas/genéticaRESUMO
Queuosine (Q) stands out as the sole tRNA modification that can be synthesized via salvage pathways. Comparative genomic analyses identified specific bacteria that showed a discrepancy between the projected Q salvage route and the predicted substrate specificities of the two identified salvage proteins: 1) the distinctive enzyme tRNA guanine-34 transglycosylase (bacterial TGT, or bTGT), responsible for inserting precursor bases into target tRNAs; and 2) Queuosine Precursor Transporter (QPTR), a transporter protein that imports Q precursors. Organisms like the facultative intracellular pathogen Bartonella henselae, which possess only bTGT and QPTR but lack predicted enzymes for converting preQ1 to Q, would be expected to salvage the queuine (q) base, mirroring the scenario for the obligate intracellular pathogen Chlamydia trachomatis. However, sequence analyses indicate that the substrate-specificity residues of their bTGTs resemble those of enzymes inserting preQ1 rather than q. Intriguingly, mass spectrometry analyses of tRNA modification profiles in B. henselae reveal trace amounts of preQ1, previously not observed in a natural context. Complementation analysis demonstrates that B. henselae bTGT and QPTR not only utilize preQ1, akin to their Escherichia coli counterparts, but can also process q when provided at elevated concentrations. The experimental and phylogenomic analyses suggest that the Q pathway in B. henselae could represent an evolutionary transition among intracellular pathogens-from ancestors that synthesized Q de novo to a state prioritizing the salvage of q. Another possibility that will require further investigations is that the insertion of preQ1 has fitness advantages when B. henselae is growing outside a mammalian host.
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Quiescent cells require a continuous supply of proteins to maintain protein homeostasis. In fission yeast, entry into quiescence is triggered by nitrogen stress, leading to the inactivation of TORC1 and the activation of TORC2. Here, we report that the Greatwall-Endosulfine-PPA/B55 pathway connects the downregulation of TORC1 with the upregulation of TORC2, resulting in the activation of Elongator-dependent tRNA modifications essential for sustaining the translation programme during entry into quiescence. This process promotes U34 and A37 tRNA modifications at the anticodon stem loop, enhancing translation efficiency and fidelity of mRNAs enriched for AAA versus AAG lysine codons. Notably, some of these mRNAs encode inhibitors of TORC1, activators of TORC2, tRNA modifiers, and proteins necessary for telomeric and subtelomeric functions. Therefore, we propose a novel mechanism by which cells respond to nitrogen stress at the level of translation, involving a coordinated interplay between the tRNA epitranscriptome and biased codon usage.