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Translation fidelity relies on accurate aminoacylation of transfer RNAs (tRNAs) by aminoacyl-tRNA synthetases (AARSs). AARSs specific for alanine (Ala), leucine (Leu), serine, and pyrrolysine do not recognize the anticodon bases. Single nucleotide anticodon variants in their cognate tRNAs can lead to mistranslation. Human genomes include both rare and more common mistranslating tRNA variants. We investigated three rare human tRNALeu variants that mis-incorporate Leu at phenylalanine or tryptophan codons. Expression of each tRNALeu anticodon variant in neuroblastoma cells caused defects in fluorescent protein production without significantly increased cytotoxicity under normal conditions or in the context of proteasome inhibition. Using tRNA sequencing and mass spectrometry we confirmed that each tRNALeu variant was expressed and generated mistranslation with Leu. To probe the flexibility of the entire genetic code towards Leu mis-incorporation, we created 64 yeast strains to express all possible tRNALeu anticodon variants in a doxycycline-inducible system. While some variants showed mild or no growth defects, many anticodon variants, enriched with G/C at positions 35 and 36, including those replacing Leu for proline, arginine, alanine, or glycine, caused dramatic reductions in growth. Differential phenotypic defects were observed for tRNALeu mutants with synonymous anticodons and for different tRNALeu isoacceptors with the same anticodon. A comparison to tRNAAla anticodon variants demonstrates that Ala mis-incorporation is more tolerable than Leu at nearly every codon. The data show that the nature of the amino acid substitution, the tRNA gene, and the anticodon are each important factors that influence the ability of cells to tolerate mistranslating tRNAs.

Biological systems are homochiral, raising the question of how a racemic mixture of prebiotically synthesized biomolecules could attain a homochiral state at the network level. Based on our recent results, we aim to address a related question of how chiral information might have flowed in a prebiotic network. Utilizing the crystallization properties of the central ribonucleic acid (RNA) precursor known as ribose-aminooxazoline (RAO), we showed that its homochiral crystals can be obtained from its fully racemic solution on a magnetic mineral surface due to the chiral-induced spin selectivity (CISS) effect [Ozturk et al., arXiv:2303.01394 (2023)]. Moreover, we uncovered a mechanism facilitated by the CISS effect through which chiral molecules, such as RAO, can uniformly magnetize such surfaces in a variety of planetary environments in a persistent manner [Ozturk et al., arXiv:2304.09095 (2023)]. All this is very tantalizing because recent experiments with tRNA analogs demonstrate high stereoselectivity in the attachment of L-amino acids to D-ribonucleotides, enabling the transfer of homochirality from RNA to peptides [Wu et al., J. Am. Chem. Soc. 143, 11836 (2021)]. Therefore, the biological homochirality problem may be reduced to ensuring that a single common RNA precursor (e.g., RAO) can be made homochiral. The emergence of homochirality at RAO then allows for the chiral information to propagate through RNA, then to peptides, and ultimately through enantioselective catalysis to metabolites. This directionality of the chiral information flow parallels that of the central dogma of molecular biology—the unidirectional transfer of genetic information from nucleic acids to proteins [F. H. Crick, in Symposia of the Society for Experimental Biology, Number XII: The Biological Replication of Macromolecules, edited by F. K. Sanders (Cambridge University Press, Cambridge, 1958), pp. 138-163; and F. Crick, Nature 227, 561 (1970)]. [ABSTRACT FROM AUTHOR]

tRNAs are evolutionarily ancient molecular decoders essential for protein translation. In eukaryotes, tRNAs and other short, noncoding RNAs are transcribed by RNA polymerase (Pol) III, an enzyme that promotes ageing in yeast, worms, and flies. Here, we show that a partial reduction in Pol III activity specifically disrupts tRNA levels. This effect is conserved across worms, flies, and mice, where computational models indicate that it impacts mRNA decoding. In all 3 species, reduced Pol III activity increases proteostatic resilience. In worms, it activates the unfolded protein response (UPR) and direct disruption of tRNA metabolism is sufficient to recapitulate this. In flies, decreasing Pol III's transcriptional initiation on tRNA genes by a loss-of-function in the TFIIIC transcription factor robustly extends lifespan, improves proteostatic resilience and recapitulates the broad-spectrum benefits to late-life health seen following partial Pol III inhibition. We provide evidence that a partial reduction in Pol III activity impacts translation, quantitatively or qualitatively, in both worms and flies, indicating a potential mode of action. Our work demonstrates a conserved and previously unappreciated role of tRNAs in animal ageing. tRNAs are important for protein translation, but whether they influence aging is unknown. In this paper, by manipulating RNA polymerase III-mediated transcription of tRNAs across three animal species, the authors demonstrate previously unappreciated roles of tRNA in proteostasis, old-age health and animal longevity. [ABSTRACT FROM AUTHOR]

'Boju' and 'Huaiju' are cultivars of the Chrysanthemum (Chrysanthemum morifolium Ramat.) in the family Asteraceae, valued for their medicinal, tea, and ornamental properties, and valued by individuals. However, the yield and quality of medicinal chrysanthemums are limited by the characteristics of the germplasm resources, including the identification at the varieties and cultivation levels. Currently, research characterizing the chloroplast genomes of medicinal Chrysanthemum flowers is relatively limited. This study conducted chloroplast whole-genome sequencing on two cultivars of Chrysanthemum, 'Boju' and 'Huaiju', and compared them with the previously published chloroplast genomes of 'Hangbaiju' and 'Gongju'. The study analyzed the chloroplast genome structures of these four medicinal Chrysanthemums, identifying mutation hotspots and clarifying their phylogenetic relationships. The chloroplast genome sizes of four medicinal Chrysanthemum cultivation products ranged from 151,057 to 151,109 bp, with GC content ranging from 37.45% to 37.76%. A total of 134 genes were identified, including 89 protein-coding genes, 37 ribosomal RNA genes, and 8 transfer RNA genes. Comparative genomic analysis revealed 159 large repeat sequences, 276 simple sequence repeats, 1 gene, and 8 intergenic regions identified as highly variable regions. Nucleotide diversity (Pi) values were high (≥ 0.004) for the petN-psbM, trnR-UCU-trnT-GGU, trnT-GGU-psbD, ndhC-trnV-UCA, ycf1, ndhI-ndhG, trnL-UGA-rpl32, rpl32-ndhF, and ndhF-ycf1 fragments, aiding in variety identification. Phylogenetic analysis revealed consistent results between maximum likelihood and Bayesian inference trees, showing that the four medicinal Chrysanthemum cultivars, along with their wild counterparts and related species, evolved as a monophyletic group, forming a sister clade to Artemisia and Ajania. Among the six Chrysanthemum species, the wild Chrysanthemum diverged first (Posterior probability = 1, bootstrap = 1,000), followed by Ajania, while C. indicum and C. morifolium clustered together (Bootstrap = 100), indicating their closest genetic relationship. The chloroplast whole-genome data and characteristic information provided in this study can be used for variety identification, genetic conservation, and phylogenetic analysis within the family Asteraceae. [ABSTRACT FROM AUTHOR]

Rampant C-to-U RNA editing drives the mutation and evolution of SARS-CoV-2. While much attention has been paid to missense mutations, the C-to-U events leading to AUG and thus creating novel ORFs were uninvestigated. By utilizing the public time-course mutation data from the worldwide SARS-CoV-2 population, we systematically identified the "AUG-gain mutations" caused by C-to-U RNA editing. Synonymous mutations were of special focus. A total of 58 synonymous C-to-U sites are able to create out-of-frame AUG in coding sequence (CDS). These 58 synonymous sites showed significantly higher allele frequency (AF) and increasing rate (dAF/dt) than other C-to-U synonymous sites in the SARS-CoV-2 population, suggesting that these 58 AUG-gain events conferred additional benefits to the virus and are subjected to positive selection. The 58 predicted new ORFs created by AUG-gain events showed the following advantages compared to random expectation: they have longer lengths, higher codon adaptation index (CAI), higher Kozak scores, and higher tRNA adaptation index (tAI). The 58 putatively novel ORFs have high expressibility and are very likely to be functional, providing an explanation for the positive selection on the 58 AUG-gain mutations. Our study proposed a possible mechanism of the emergence of de novo genes in SARS-CoV-2. This idea should be helpful in studying the mutation and evolution of SARS-CoV-2. [ABSTRACT FROM AUTHOR]

Nonuniform local translation speed dictates diverse protein biogenesis outcomes. To unify known and uncover unknown principles governing eukaryotic elongation rate, we developed a machine learning pipeline to analyze RiboSeq datasets. We find that the chemical nature of the incoming amino acid determines how codon optimality influences elongation rate, with hydrophobic residues more dependent on transfer RNA (tRNA) levels than charged residues. Unexpectedly, we find that wobble interactions exert a widespread effect on elongation pausing, with wobble-mediated decoding being slower than Watson-Crick decoding, irrespective of tRNA levels. Applying our ribosome pausing principles to ribosome collisions reveals that disomes arise upon apposition of fast-decoding and slow-decoding signatures. We conclude that codon choice and tRNA pools are evolutionarily constrained to harmonize elongation rate with cotranslational folding while minimizing wobble pairing and deleterious stalling. [ABSTRACT FROM AUTHOR]

Background and Purpose Experimental Approach Key Results Conclusions and Implications Renal interstitial fibrosis is a frequent pathological manifestation of lupus nephritis (LN). tRNA halves (tiRNAs) are acquired from tRNA‐derived small non‐coding RNAs (sncRNAs) and are associated with fibrosis. Our previous study indicated enhanced tiRNA‐Gly‐GCC‐002 (tiRNA002) levels in kidneys were positively related to LN‐related fibrosis. However, the precise molecular mechanism remains unclear.The mimic and agomiR of tiRNA002 were introduced into tubular epithelial cells (TECs) and MRL/lpr mice by transfection. The levels of gene and protein expressions were quantified using real‐time quantitative polymerase chain reaction (RT‐qPCR), Western blot and immunofluorescence assays.In TECs treated with LN serum, as well as in the kidneys of MRL/lpr mice, high levels of tiRNA002 directly influenced the epithelial‐mesenchymal transition (EMT) and extracellular matrix (ECM) deposition. Furthermore, tiRNA002 overexpression promoted EMT in TECs and accelerated renal interstitial fibrosis in MRL/lpr mice via Smad signalling. The target gene of tiRNA002, FKBP prolyl isomerase 5 (FKBP5), improved Smad signalling by interacting with phosphorylated Smad2/3. Silencing FKBP5 alleviated LN serum‐ or tiRNA002‐mimic‐induced EMT in TECs. In addition, FKBP5 overexpression reversed the tiRNA002 knockdown‐mediated reduction of EMT and ECM accumulation.These findings indicated that tiRNA002 is markedly increased in LN, which facilitates renal fibrosis by promoting EMT via FKBP5‐mediated Smad signalling. Therefore, targeting tiRNA002 may be an innovative approach to treat renal interstitial fibrosis in LN. [ABSTRACT FROM AUTHOR]

Aminoacyl‐tRNA synthetases (aaRSs) are universally essential enzymes that synthesize aminoacyl‐tRNA substrates for protein synthesis. Although most organisms require a single aaRS gene for each proteinogenic amino acid to translate their genetic information, numerous species encode multiple gene copies of an aaRS. Growing evidence indicates that organisms acquire extra aaRS genes to sustain or adapt to their unique lifestyle. However, predicting and defining the function of repeated aaRS genes remains challenging due to their potentially unique physiological role in the host organism and the inconsistent annotation of repeated aaRS genes in the literature. Here, we carried out comparative, phylogenetic, and functional studies to determine the activity of coexisting paralogs of tryptophanyl‐, tyrosyl‐, seryl‐, and prolyl‐tRNA synthetases encoded in several human pathogenic bacteria. Our analyses revealed that, with a few exceptions, repeated aaRSs involve paralogous genes with distinct phylogenetic backgrounds. Using a collection of Escherichia coli strains that enabled the facile characterization of aaRS activity in vivo, we found that, in almost all cases, one aaRS displayed transfer RNA (tRNA) aminoacylation activity, whereas the other was not compatible with E. coli. Together, this work illustrates the challenges of identifying, classifying, and predicting the function of aaRS paralogs and highlights the complexity of aaRS evolution. Moreover, these results provide new insights into the potential role of aaRS paralogs in the biology of several human pathogens and foundational knowledge for the investigation of the physiological role of repeated aaRS paralogs across bacteria. [ABSTRACT FROM AUTHOR]

5-Methylcytosine (5mC) is an established epigenetic mark in vertebrate genomic DNA, but whether its oxidation intermediates formed during TET-mediated DNA demethylation possess an instructive role of their own that is also physiologically relevant remains unresolved. Here, we reveal a 5-formylcytosine (5fC) nuclear chromocenter, which transiently forms during zygotic genome activation (ZGA) in Xenopus and mouse embryos. We identify this chromocenter as the perinucleolar compartment, a structure associated with RNA Pol III transcription. In Xenopus embryos, 5fC is highly enriched on Pol III target genes activated at ZGA, notably at oocyte-type tandem arrayed tRNA genes. By manipulating Tet and Tdg enzymes, we show that 5fC is required as a regulatory mark to promote Pol III recruitment as well as tRNA expression. Concordantly, 5fC modification of a tRNA transgene enhances its expression in vivo. The results establish 5fC as an activating epigenetic mark during zygotic reprogramming of Pol III gene expression. [Display omitted] • 5fC forms chromocenters in the perinucleolar compartment during Xenopus ZGA • 5fC colocalizes with RNA Pol III and accumulates on active tRNA tandem repeats • 5fC promotes Pol III recruitment and tRNA transcription at ZGA • 5fC stimulates transcription of a 5fC-modified tRNA transgene An open question in epigenetics is if 5-formylcytosine (5fC) is merely a byproduct of active demethylation or an instructive mark that regulates gene expression on its own. This work demonstrates 5fC chromocenters in early frog embryos and shows that 5fC acts as an instructive mark to promote RNA Pol III transcription during zygotic genome activation. [ABSTRACT FROM AUTHOR]

Human mitochondrial tRNAs (mt-tRNAs), critical for mitochondrial biogenesis, are frequently associated with pathogenic mutations. These mt-tRNAs have unusual sequence motifs and require post-transcriptional modifications to stabilize their fragile structures. However, whether a modification that stabilizes a wild-type (WT) mt-tRNA would also stabilize its pathogenic variants is unknown. Here we show that the N1-methylation of guanosine at position 9 (m1G9) of mt-Leu(UAA), while stabilizing the WT tRNA, has a destabilizing effect on variants associated with MELAS (mitochondrial myopathy, encephalopathy, lactic acidosis, and stroke-like episodes). This differential effect is further demonstrated, as removal of the m1G9 methylation, while damaging to the WT tRNA, is beneficial to the major pathogenic variant, improving the structure and activity of the variant. These results have therapeutic implications, suggesting that the N1-methylation of mt-tRNAs at position 9 is a determinant of pathogenicity and that controlling the methylation level is an important modulator of mt-tRNA-associated diseases. Here the authors show that the m1G9 post-transcriptional methylation differentially regulates the stability of the native and the MELAS variants of human mt-Leu(UAA) tRNA, contributing to mitochondrial pathology. [ABSTRACT FROM AUTHOR]

The quantity of each protein in a cell only is only partially correlated with its gene transcription rate. Independent influences on protein synthesis levels include mRNA sequence motifs, amino acyl-tRNA synthesis levels, elongation factor action, and protein susceptibility to degradation. Here we report that the amino acid composition of a protein can also influence its expression level in two distinct ways. The nutritional classification of amino acids in animals reflects their potential for scarcity–essential amino acids (EAA) are reliant on dietary supply, non-essential amino acids (NEAA) from internal biosynthesis, and conditionally essential amino acids (CEAA) from both. Accessing public proteomic datasets, we demonstrate that a protein's CEAA sequence composition is inversely correlated with expression–a correlation enhanced during rapid cellular proliferation–suggesting CEAA availability can limit translation. Similarly, proteins with the most extreme compositions of EAA are generally reduced in abundance. These latter proteins participate in biological systems such as taste and food-seeking behaviour, oxidative phosphorylation, and chemokine function, and so linking their expression to EAA availability may act as a homeostatic response to malnutrition. Protein composition can also influence general human phenotypes and disease susceptibility: stature proteins are enriched in CEAAs, and a curated dataset of over 700 cancer proteins is significantly under-represented in EAAs. We also show that individual amino acids can influence protein expression across all kingdoms of life and that this effect appears to be rooted in the unchanging structural and mRNA encoding features of each amino acid. Species-specific environmental survival pathways are shown to be enriched in proteins with individual amino acid compositions favouring higher expression. These two forms of amino acid-driven protein expression regulation promise new insights into systems biology, evolutionary studies, experimental research design, and public health intervention. [ABSTRACT FROM AUTHOR]

Background: Archaea and Bacteria are distinct domains of life that are adapted to a variety of ecological niches. Several genome-based methods have been developed for their accurate classification, yet many aspects of the specific genomic features that determine these differences are not fully understood. In this study, we used publicly available whole-genome sequences from bacteria ( N = 2546 ) and archaea ( N = 109 ). From these, a set of genomic features (nucleotide frequencies and proportions, coding sequences (CDS), non-coding, ribosomal and transfer RNA genes (ncRNA, rRNA, tRNA), Chargaff's, topological entropy and Shannon's entropy scores) was extracted and used as input data to develop machine learning models for the classification of archaea and bacteria. Results: The classification accuracy ranged from 0.993 (Random Forest) to 0.998 (Neural Networks). Over the four models, only 11 examples were misclassified, especially those belonging to the minority class (Archaea). From variable importance, tRNA topological and Shannon's entropy, nucleotide frequencies in tRNA, rRNA and ncRNA, CDS, tRNA and rRNA Chargaff's scores have emerged as the top discriminating factors. In particular, tRNA entropy (both topological and Shannon's) was the most important genomic feature for classification, pointing at the complex interactions between the genetic code, tRNAs and the translational machinery. Conclusions: tRNA, rRNA and ncRNA genes emerged as the key genomic elements that underpin the classification of archaea and bacteria. In particular, higher nucleotide diversity was found in tRNA from bacteria compared to archaea. The analysis of the few classification errors reflects the complex phylogenetic relationships between bacteria, archaea and eukaryotes. [ABSTRACT FROM AUTHOR]

Epinephelus bilobatus, Epinephelus maculatus and Epinephelus longispinis are three groupers that share common morphological characteristics and coloration patterns and have been morphologically confused and misidentified with each other for a long time. Complete mitochondrial genomes of the three groupers were determined and analyzed in this study. Mitogenomes of E. bilobatus, E. maculatus and E. longispinis were 17, 354 bp, 17, 066 bp and 17, 221 bp in size respectively and consisted of 13 protein-coding genes, two ribosomal RNA genes and one control region. However, different from most teleosts, which contain canonical 22 tRNAs, more numbers of tRNAs were identified in the three groupers with 27 tRNAs in E. bilobatus and E. longispinis and 25 tRNAs in E. maculatus. The increased number of tRNAs was due to the presence of tandemly duplicated tRNA-Asp genes that were located between tRNA-Ser and COII genes (six duplications in E. bilobatus and E. longispinis, four duplications in E. maculatus). Intact gene tandem duplication was an uncommon feature that was found in the typical teleost mitogenomes. The phylogenetic trees of the 32 groupers (genus Epinephelus) that were constructed based on 12 protein-coding genes revealed that Epinephelus species with tandemly duplicated tRNA-Asp genes were clustered into one monophyletic group, distinct from other Epinephelus species without any duplication features, which indicated that tandemly duplicated tRNA-Asp genes may be the particular linage-specific characteristics that evolve from a common ancestor and have the ability to distinguish them from other Epinephelus species. The results of the mitogenomes comparative analyses of the three groupers revealed the genetic distance of mitogenomes between each two species to be 0.062 (E. bilobatus vs E. maculatus), 0.091 (E. bilobatus vs E. longispinis) and 0.087 (E. maculatus vs E. longispinis). All values were far greater than the minimum value of 0.020 for species identification, which shows that they were three independent species at molecular level. Regarding the relationships between the three groupers, E. bilobatus was found to be more closely related to E. maculatus in comparison to E. longispinis. The results provide valuable molecular data for the species identification and phylogenetic analyses on E. bilobatus, E. maculatus and E. longispinis, and also provided a new insight into the tandem gene duplication features of Epinephelus mitogenomes. [ABSTRACT FROM AUTHOR]

Small non-coding RNAs (sncRNAs) derived from tRNAs are known as tRNA-derived small RNAs (tsRNAs). These tsRNAs are further categorized into tRNA-derived fragments (tRFs) and tRNA halves (tiRNAs), which play significant roles in the various molecular mechanisms underlying certain human diseases. However, the generation of tsRNAs and their potential roles during Dengue virus (DENV) infection is not yet known. Here, we performed small RNA sequencing to identify the generation and alterations in tsRNAs expression profiles of DENV-infected Huh7 cells. Upon DENV infection, tRNA fragmentation was found to be increased. We identified a significant number of differentially expressed tsRNAs during DENV infection. Interestingly, the 3′tRF population showed upregulation, while the i-tRF population exhibited downregulation. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis was performed to analyze the impact of differentially expressed tsRNAs on DENV pathogenesis. Our results suggest that differentially expressed tsRNAs are involved in transcriptional regulation via RNA polymerase II promoter and metabolic pathways. Overall, our study contributes significantly to our understanding of the roles played by tsRNAs in the complex dynamics of DENV infection. [ABSTRACT FROM AUTHOR]

Intrauterine growth restriction (IUGR) impairs neonatal weight and causes multiple organ dysplasia. IUGR not only threatens human health but is also a significant constraint to the development of animal husbandry. However, the molecular mechanism underlying IUGR remains to be further elucidated. tRNA-derived small RNA (tsRNAs) is a regulative non-coding RNA, which has recently been reported to correlate with the onset and progression of several diseases. In this study, we investigated the tsRNAs expression profiles of IUGR pigs. A tsRNAs dataset for multiple organs in normal and IUGR pigs was generated, including muscle, liver, spleen and intestine. We further analyzed the characteristics of tsRNAs in different organs of pigs, and KEGG pathway analysis was performed to investigate possible pathways involved. This dataset will provide valuable information for further exploring the molecular mechanism of IUGR formation. [ABSTRACT FROM AUTHOR]

The succulent shrubs Desmidorchis penicillata and D. retrospiciens, part of the taxonomically challenging genus Desmidorchis, are well‐known for their ecological resilience and medicinal significance. This study sequences the first complete chloroplast genomes of these species, shedding light on their genomic characteristics and evolutionary relationships. The circular genomes of D. penicillata (161 776 bp) and D. retrospiciens (162 277 bp) display a quadripartite structure typical of Angiosperms. Gene content, order, and GC content are consistent, featuring 114 unique genes, including 80 protein‐coding, 30 transfer RNAs, and four ribosomal RNAs genes. Codon usage analysis underscores A/U‐rich preferences, while RNA editing sites, predominantly in ndhB and ndhD genes, suggest post‐transcriptional modifications. Analysis of long repeated sequences reveals a predominance of forward and palindromic repeats. Simple sequence repeats (SSRs), particularly A/T motifs, are abundant, with high presence of mononucleotide, offering potential molecular markers. Comparative analysis with their relatives in subtribe Stapeliinae identifies mutational hotspots such as ycf1, ndhF, trnG(GCC)‐trnfM(CGA) and ndhG‐ndhI that could be potential DNA barcoding markers. The inverted repeat (IR) boundaries analysis revealed an expansion of IR on the small single copy region, leading to the formation of a pseudogene. Overall, substitution rate analysis indicated purifying selection, with a few genes (rpl22, clpP and rps11) showing signatures of positive selection. Additionally, the phylogenetic analysis positioned Desmidorchis within the Stapeliinae clade and strongly supported the sister relationship between D. penicillata and D. retrospiciens. This study provides comprehensive molecular data for future research in Desmidorchis. [ABSTRACT FROM AUTHOR]

Aminoacyl-tRNA synthetases hold the key to the genetic code and assign nucleic acid-based codons to amino acids, the building blocks of proteins. In their ability to recognize identity elements on transfer RNAs (tRNAs), some as simple as a single base pair, they ensure that the same proteins are formed each time information embedded in DNA is transcribed into messenger RNA (mRNA) and translated into proteins (Figure 1A). Thus, aminoacyl-tRNA synthetase active sites are conserved; however, since their evolutionary origin, their functions have been co-opted, expanded on and played novel roles during evolution. Below, we provide an overview of the many functions of aminoacyl-tRNA synthetases — from their role in translation, one of the most fundamental processes of all life, to newly discovered, diverse functions. Rasangi Tennakoon and Haissi Cui introduce aminoacyl-tRNA synthetases, the class of enzymes that load amino acids onto tRNAs and thus interpret the genetic code. [ABSTRACT FROM AUTHOR]

Two new species of Dulcicalothrix, D. adhikaryi sp. nov. and D. iyengarii sp. nov., were discovered in India and are characterized and described in accordance with the rules of the International Code of Nomenclature for algae, fungi, and plants (ICN). As a result of phylogenetic analysis, Calothrix elsteri is reassigned to Brunnivagina gen. nov. During comparison with all Dulcicalothrix for which sequence data were available, we observed that the genus has six ribosomal operons in three orthologous types. Each of the three orthologs could be identified based upon indels occurring in the D1–D1′ helix sequence in the ITS rRNA region between the 16S and 23S rRNA genes, and in these three types, there were operons containing ITS rRNA regions with and without tRNA genes. Examination of complete genomes in Dulcicalothrix revealed that, at least in the three strains for which complete genomes are available, there are five ribosomal operons, two with tRNA genes and three with no tRNA genes in the ITS rRNA region. Internal transcribed spacer rRNA regions have been consistently used to differentiate species, both on the basis of secondary structure and percent dissimilarity. Our findings call into question the use of ITS rRNA regions to differentiate species in the absence of efforts to obtain multiple operons of the ITS rRNA region through cloning or targeted PCR amplicons. The ITS rRNA region data for Dulcicalothrix is woefully incomplete, but we provide herein a means for dealing with incomplete data using the polyphasic approach to analyze diverse molecular character sets. Caution is urged in using ITS rRNA data, but a way forward through the complexity is also proposed. [ABSTRACT FROM AUTHOR]

Brachidontes variabilis (Krauss, 1848) is a common small mussel widely distributed in the intertidal zone of southern China. Firstly, we characterized four complete mitochondrial genomes of B. variabilis from four populations (Wanning, Yangjiang, Zhangzhou, and Shanwei) in Chinese waters. This study was the first to sequence and discribe the mitochondrial genome of B. variabilis, which comprised of 2 ribosomal RNAs (rRNAs), 22 transfer RNAs (tRNAs) and 12 protein-coding genes (PCGs, lacking atp8). And then three PCGs fragments, including cytochrome oxidase subunit I (cox1, 558 bp), NADH dehydrogenase subunit 5 (nad5, 754 bp), and NADH dehydrogenase subunit 4 (nad4, 879 bp), were utilized to investigate the population genetics and demography. The cox1, nad5 and nad4 fragments revealed 55, 81, and 103 haplotypes, respectively. Results indicated high genetic diversity within populations (Hd > 0.5, Pi > 0.005) and little genetic differentiation among populations (FST<0.05), suggesting frequent gene exchanges. Further, the neutrality test and Extended Bayesian skyline plots (EBSPs) suggested that the four populations underwent population expansion at 0.9–1.1 Ma during the Pleistocene. In summary, this study on the mitochondrial genomes and population genetics of B. variabilis provides insights into the evolutionary history and demographic dynamics of this species in southern China. [ABSTRACT FROM AUTHOR]

Simple Summary: The present study aimed to assemble and annotate the mitochondrial genome of Keenocardium buelowi and conducted an in-depth analysis to investigate the molecular features of this mitochondrial genome. Our findings established phylogenetic relationships within the Cardiidae family. Additionally, evidence of selection pressure on the cytochrome b gene was detected, suggesting its important role in the evolution of K. buelowi. The mitochondrial genome provides valuable data for phylogenetic analysis and evolutionary research. In this study, we sequenced, assembled, and annotated the mitochondrial genome of Keenocardium buelowi using the Illumina platform. The genome spanned 16,967 bp and included 13 protein-coding genes (PCGs), two ribosomal RNAs, and 22 transfer RNAs. All PCGs utilized standard ATN start codons and TAN stop codons. The phylogenetic tree based on maximum likelihood and Bayesian inference analyses revealed Clinocardiinae as the sister group to Trachycardiinae, with the estimated divergence time being 44.5 million years ago (MYA) between K. buelowi and Vasticardium flavum. Notably, the cytochrome b gene (cob) exhibited a positive selection signal. Our findings provide valuable insights into the evolutionary history and molecular phylogeny of K. buelowi. [ABSTRACT FROM AUTHOR]

The loss of one of the two copies of the 9 bp tandem repeat sequence (CCCCCTCTA) located in the small non-coding region between the cytochrome oxidase II (COII) and the lysine tRNA genes in human mtDNA has been reported to be polymorphic in Asian, Oceanian and Sub-Saharan African populations, but it has rarely been observed in Europe. In this study, we will evaluate the possible association between the MIC9D polymorphism and cognitive disorders. A genetic analysis of unrelated Sicilian patients with cognitive deficits was performed to identify the 9 bp deletion MIC9D polymorphism. The MIC9D polymorphism was found in six patients, whereas this variant was absent in control individuals without cognitive deficits. The patients with the MIC9D polymorphism exhibited more complex clinical presentations; in particular, all had neuromuscular disorders and five also presented with behavioral disorders. The present study suggests a potential association between the MIC9D polymorphism and cognitive impairment with concurrent neuromuscular and behavioral involvement. [ABSTRACT FROM AUTHOR]

The evolutionary history of emperors, particularly in the Atlantic and Indo-West Pacific Oceans, remains largely unmapped. This study explores the maternal lineage evolution of Lethrinids by examining the complete mitogenome of Lethrinus atlanticus, which is endemic to the Eastern Atlantic Ocean. Utilizing advanced next-generation sequencing, we found that the mitogenome spans 16,789 base pairs and encompasses 37 genes, including 13 protein-coding genes (PCGs), two ribosomal RNAs, 22 transfer RNAs, and an AT-rich control region (CR). Our analysis indicates a preference for AT base pairs in the L. atlanticus mitogenome (53.10%). Most PCGs begin with the ATG codon, except for COI, which starts with GTG. Relative synonymous codon usage reveals high frequencies for alanine, leucine, proline, serine, and threonine. The ratio of nonsynonymous to synonymous substitutions suggests strong negative selection across all PCGs in Lethrinus species. Most transfer RNAs exhibit typical cloverleaf structures, with the exception of tRNA-serine (GCT), which lacks a dihydrouracil stem. Comparative analysis of conserved sequence blocks across the CRs of three Lethrinus species shows notable differences in length and nucleotide composition. Phylogenetic analysis using concatenated PCGs clearly distinguishes all Lethrinus species, including L. atlanticus, and sheds light on the evolutionary relationships among Spariformes species. The estimated divergence time of approximately 20.67 million years between L. atlanticus and its Indo-West Pacific relatives provides insights into their historical separation and colonization during the late Oligocene. The distribution of Lethrinids may be influenced by ocean currents and ecological factors, potentially leading to their speciation across the Eastern Atlantic and Indo-West Pacific. This study enhances our understanding of the genetic diversity and phylogenetic relationships within Lethrinus species. Further exploration of other emperor fish mitogenomes and comprehensive genomic data could provide vital insights into their genetic makeup, evolutionary history, and environmental adaptability in marine ecosystems globally. [ABSTRACT FROM AUTHOR]

Retrotransposons are mobile DNA elements that are more active with increasing age and exacerbate aging phenotypes in multiple species. We previously reported an unexpected extension of chronological lifespan in the yeast, Saccharomyces paradoxus, due to the presence of Ty1 retrotransposons when cells were aged under conditions of mild stress. In this study, we tested a subset of genes identified by RNA-seq to be differentially expressed in S. paradoxus strains with a high-copy number of Ty1 retrotransposons compared with a strain with no retrotransposons and additional candidate genes for their contribution to lifespan extension when cells were exposed to a moderate dose of hydroxyurea (HU). Deletion of ADE8, NCS2, or TRM9 prevented lifespan extension, while deletion of CDD1, HAC1, or IRE1 partially prevented lifespan extension. Genes overexpressed in high-copy Ty1 strains did not typically have Ty1 insertions in their promoter regions. We found that silencing genomic copies of Ty1 prevented lifespan extension, while expression of Ty1 from a high-copy plasmid extended lifespan in medium with HU or synthetic medium. These results indicate that cells adapt to expression of retrotransposons by changing gene expression in a manner that can better prepare them to remain healthy under mild stress. [ABSTRACT FROM AUTHOR]

Sugarcane thrips, Fulmekiola serrata (Kobus) (Thysanoptera: Thripidae), is a common foliar pest that infests sugarcane and is found throughout tropical and subtropical countries. In this study, we obtained and analyzed the complete mitochondrial genome of F. serrata for the first time and explored the phylogenetic relationships of the higher-order elements of Thysanoptera members at the mitochondrial level. The complete mitochondrial genome of F. serrata is 16,596 bp in length and includes 13 protein-coding genes (PCGs), 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes, and 1 noncoding control region. A+T accounted for 75% of the total bases in the mitochondrial genome of F. serrata, revealing an obvious AT bias. Among the 13 PCGs, except for nad5, which had a start codon of TTG, the remaining genes had ATNs typical of insects (ATA, ATT, ATC, and ATG); nad1, nad2, nad3, and atp8 had incomplete termination codons of TA or T. The remaining nine PCGs were complete with the termination codon TAA. Of the 22 tRNA secondary structures, all were typical cloverleaf secondary structures except for trnS1, which was missing the DHU arm. Compared with the hypothetical ancestral gene arrangement of arthropods, F. serrata presented extensive gene rearrangement, with 23 translocated genes, 8 inverted genes, and 5 shuffled genes. Both maximum likelihood (ML) and Bayesian inference (BI) phylogenetic trees resulted in similar topologies: ((Thripidae + (Stenurothripidae + Aeolothripidae)) + Phlaeothripidae), with Thripidae, Aeolothripidae and Phlaeothripidae being monophyletic groups, whereas F. serrata is closely related to Thrips palmi, and the two are sister groups. [ABSTRACT FROM AUTHOR]

Heterozygous pathogenic variants in the histidyl‐tRNA synthetase (HARS) gene are associated with Charcot–Marie–Tooth (CMT) type 2W disease, classified as an axonal peripheral neuropathy. To date, at least 60 variants causing CMT symptoms have been identified in seven different aminoacyl‐tRNA synthetases, with eight being found in the catalytic domain of HARS. The genetic data clearly show a causative role of aminoacyl‐tRNA synthetases in CMT; however, the cellular mechanisms leading to pathology can vary widely and are unknown in the case of most identified variants. Here we describe a novel HARS variant, c.412T>C; p.Y138H, identified through a CMT gene panel in a patient with peripheral neuropathy. To determine the effect of p.Y138H we employed a humanized HARS yeast model and recombinant protein biochemistry, which identified a deficiency in protein dimerization and a growth defect which shows mild but significant improvement with histidine supplementation. This raises the potential for a clinical trial of histidine. [ABSTRACT FROM AUTHOR]

Codon optimality refers to the effect that codon composition has on messenger RNA (mRNA) stability and translation level and implies that synonymous codons are not silent from a regulatory point of view. Here, we investigated the adaptation of virus genomes to the host optimality code using mosquito-borne dengue virus (DENV) as a model. We demonstrated that codon optimality exists in mosquito cells and showed that DENV preferentially uses nonoptimal (destabilizing) codons and avoids codons that are defined as optimal (stabilizing) in either human or mosquito cells. Human genes enriched in the codons preferentially and frequently used by DENV are upregulated during infection, and so is the tRNA decoding the nonoptimal and DENV preferentially used codon for arginine. We found that adaptation during single-host passaging in human or mosquito cells results in the selection of synonymous mutations towards DENV's preferred nonoptimal codons that increase virus fitness. Finally, our analyses revealed that hundreds of viruses preferentially use nonoptimal codons, with those infecting a single host displaying an even stronger bias, suggesting that host–pathogen interaction shapes virus-synonymous codon choice. Synopsis: Analyses of viral genome adaptation to the host optimality code, reveal that hundreds of human-infecting viruses preferentially use non-optimal codons and suggest that host-pathogen interactions can shape virus synonymous codon choice. Dengue virus (DENV) preferentially uses non-optimal codons and avoids codons that are defined as optimal in either human or mosquito cells. Codon optimality mechanism exists in mosquito cells. Human genes enriched in the codons preferentially and frequently used by DENV are up-regulated during DENV infection, as well as tRNAs decoding the non-optimal codon that DENV preferentially uses. Hundreds of human-infecting viruses preferentially use non-optimal codons suggesting that host-pathogen interaction shapes virus synonymous codon choice. Analyses of viral genome adaptation to the host optimality code, reveal that hundreds of human-infecting viruses preferentially use non-optimal codons and suggest that host-pathogen interactions can shape virus synonymous codon choice. [ABSTRACT FROM AUTHOR]

Human cytoplasmic tRNAs contain dihydrouridine modifications at positions 16 and 17 (D16/D17). The enzyme responsible for D16/D17 formation and its cellular roles remain elusive. Here, we identify DUS1L as the human tRNA D16/D17 writer. DUS1L knockout in the glioblastoma cell lines LNZ308 and U87 causes loss of D16/D17. D formation is reconstituted in vitro using recombinant DUS1L in the presence of NADPH or NADH. DUS1L knockout/overexpression in LNZ308 cells shows that DUS1L supports cell growth. Moreover, higher DUS1L expression in glioma patients is associated with poorer prognosis. Upon vector-mediated DUS1L overexpression in LNZ308 cells, 5′ and 3′ processing of precursor tRNATyr(GUA) is inhibited, resulting in a reduced mature tRNATyr(GUA) level, reduced translation of the tyrosine codons UAC and UAU, and reduced translational readthrough of the near-cognate stop codons UAA and UAG. Moreover, DUS1L overexpression increases the amounts of several D16/D17-containing tRNAs and total cellular translation. Our study identifies a human dihydrouridine writer, providing the foundation to study its roles in health and disease. DUS1L was identified as the human tRNA dihydrouridine writer for positions 16 and 17. DUS1L overexpression changes the amount of several D16/D17-containing tRNAs and increases total cellular translation and cell growth in glioblastoma cells. [ABSTRACT FROM AUTHOR]

Meropenem is commonly used against Pseudomonas aeruginosa. Traditionally, the time unbound antibiotic concentration exceeds the MIC (f T > MIC) is used to select carbapenem regimens. We aimed to characterize the effects of different baseline resistance mechanisms on bacterial killing and resistance emergence; evaluate whether f T > MIC can predict these effects; and, develop a novel Quantitative and Systems Pharmacology (QSP) model to describe the effects of baseline resistance mechanisms on the time-course of bacterial response. Seven isogenic P. aeruginosa strains with a range of resistance mechanisms and MICs were used in 10-day hollow-fiber infection model studies. Meropenem pharmacokinetic profiles were simulated for various regimens (t 1/2,meropenem = 1.5 h). All viable counts on drug-free, 3 × MIC, and 5 × MIC meropenem-containing agar across all strains, five regimens, and control (n = 90 profiles) were simultaneously subjected to QSP modeling. Whole genome sequencing was completed for total population samples and emergent resistant colonies at 239 h. Regimens achieving ≥98% f T >1 × MIC suppressed resistance emergence of the mexR knockout strain. Even 100% f T >5 × MIC failed to achieve this against the strain with OprD loss and the ampD and mexR double-knockout strain. Baseline resistance mechanisms affected bacterial outcomes, even for strains with the same MIC. Genomic analysis revealed that pre-existing resistant subpopulations drove resistance emergence. During meropenem exposure, mutations in mexR were selected in strains with baseline oprD mutations, and vice versa , confirming these as major mechanisms of resistance emergence. Secondary mutations occurred in lysS or argS , coding for lysyl and arginyl tRNA synthetases, respectively. The QSP model well-characterized all bacterial outcomes of the seven strains simultaneously, which f T > MIC could not. [Display omitted] [ABSTRACT FROM AUTHOR]

Objective: This study aimed to identify, discover and explore the characteristics of the mtDNA genomes of Cemani chicken (Gallus gallus). Methods: This study used gDNA of Cemani chicken isolated from liver tissue. mtDNA sequencing was performed using WGS mtDNA analysis with nanopore technology by Oxford Nanopore Technologies GridION. Bioinformatics and data analysis were then performed. Results: This study showed that the length of the mtDNA genome is 16,789 bp, consisting of two ribosomal RNA (12S rRNA, 16S rRNA), 22 transfer RNA genes (trnR, trnG, trnK, trnD, trnS, trnY, trnC, trnN, trnA, trnW, trnM, trnQ, trnl, trnL, trnV, trnF, trnP, trnT, trnE, trnL, trnS, trnH), 13 protein-coding genes (PCGs) (ND4l, ND3, COX3, ATP6, ATP8, COX2, COX1, ND2, ND1, CYTB, ND6, ND5, ND4), and a noncoding control region (Dloop). Furthermore, analysis showed there were polymorphic sites and amino acid alterations when mtDNA Cemani chicken was aligned with references from GenBank. Conclusion: Site (988T>*) in Dloop genes and (328A>G) in ND3 genes which alter glycine to stop codon, were specific markers found only in Cemani chicken. [ABSTRACT FROM AUTHOR]

RNA interference (RNAi) shows great potential in plant defense against pathogens through RNA-mediated sequence-specific gene silencing. Among RNAi-based plant protection strategies, spray-induced gene silencing (SIGS) is considered a more promising approach because it utilizes the transfer of exogenous RNA between plants and microbes to silence target pathogen genes. The application of nanovesicles significantly enhances RNA stability and delivery efficiency, thereby improving the effectiveness of SIGS and further enhancing plant resistance to diseases and pathogens. This review explores the role of RNAi in plant protection, focusing on the cross-kingdom transport of small RNAs (sRNAs) via extracellular vesicles. It also explores the potential of nanotechnology to further optimize RNA-based plant protection, offering innovative tools and methods in modern plant biotechnology. [ABSTRACT FROM AUTHOR]

Suppressor tRNAs, notable for their capability of reading through the stop codon while maintaining normal peptide synthesis, are promising in treating diseases caused by premature termination codons (PTC). However, the lack of effective engineering methods for suppressor tRNAs has curtailed their application potential. Here, we introduce a directed evolution technology that employs phage-assisted continuous evolution (PACE), combined with gradient biosensors featuring various PTCs in the M13 gene III. Utilizing this novel methodology, we have successfully evolved tRNATrp (UGG) reading through the UGA stop codon in Escherichia coli. Massively parallel sequencing revealed that these mutations predominantly occurred in the anticodon loop. Finally, two suppressor tRNATrp (UGA) mutants exhibited over fivefold increases in readthrough efficiency. [Display omitted] • A directed evolution technology for suppressor tRNAs was developed in E. coli. • Suppressor tRNA mutations mainly occurred in the anticodon loop. • Two suppressor tRNATrp (UGA) exhibited over fivefold increase in efficiency. [ABSTRACT FROM AUTHOR]

Modification of guanosine to N7-methylguanosine (m7G) in the variable loop region of tRNA is catalyzed by the METTL1/WDR4 heterodimer and stabilizes target tRNA. Here, we reveal essential functions of Mettl1 in Drosophila fertility. Knockout of Mettl1 (Mettl1-KO) causes no major effect on the development of non-gonadal tissues, but abolishes the production of elongated spermatids and mature sperm, which is fully rescued by expression of a Mettl1-transgene, but not a catalytic-dead Mettl1 transgene. This demonstrates that Mettl1-dependent m7G is required for spermatogenesis. Mettl1-KO results in a loss of m7G modification on a subset of tRNAs and decreased tRNA abundance. Ribosome profiling shows that Mettl1-KO led to ribosomes stalling at codons decoded by tRNAs that were reduced in abundance. Mettl1-KO also significantly reduces the translation efficiency of genes involved in elongated spermatid formation and sperm stability. Germ cell-specific expression of Mettl1 rescues disrupted m7G tRNA modification and tRNA abundance in Mettl1-KO testes but not in non-gonadal tissues. Ribosome stalling is much less detectable in non-gonadal tissues than in Mettl1-KO testes. These findings reveal a developmental role for m7G tRNA modification and indicate that m7G modification-dependent tRNA abundance differs among tissues. Here the authors reveal essential functions of Mettl1, catalyzing N7-guanosine methylation (m7G) of tRNAs, in Drosophila male fertility through a regulation of the steady-state level of tRNAs and the translational efficiency of genes essential for spermatogenesis. [ABSTRACT FROM AUTHOR]

tRNAs have recently gained attention for their novel regulatory roles in translation and for their diverse functions beyond translation. One of the most remarkable aspects of tRNA biogenesis is the incorporation of various chemical modifications, ranging from simple base or ribose methylation to more complex hypermodifications such as formation of queuosine and wybutosine. Some tRNAs are transcribed as intron-containing pre-tRNAs. While the majority of these modifications occur independently of introns, some are catalyzed in an intron-inhibitory manner, and in certain cases, they occur in an intron-dependent manner. This review focuses on pre-tRNA modification, including introncontaining pre-tRNA, in both intron-inhibitory and intron-dependent fashions. Any perturbations in the modification and processing of tRNAs may lead to a range of diseases and disorders, highlighting the importance of understanding these mechanisms in molecular biology and medicine. [ABSTRACT FROM AUTHOR]

The Siberian Scoter (Melanitta stejnegeri) is a medium sea duck distinct from M. deglandi due to the absence of hybridization and differences in morphological characteristics. However, knowledge of its phylogenetic relationships within Anseriformes is limited due to a lack of molecular data. In this study, the complete mitogenome of M. stejnegeri was firstly sequenced, then annotated and used to reconstruct the phylogenetic relationships of 76 Anseriformes species. The complete mitogenome of M. stejnegeri is 16,631 bp and encodes 37 typical genes: 13 protein-coding genes, 2 ribosomal RNAs, 22 transfer RNAs, and 1 non-coding control region. Its mitogenome organization is similar to that of other Anseriformes species. The phylogenetic relationships within the genus Melanitta are initially clarified, with M. americana at the base. M. stejnegeri and M. deglandi are sister groups, clustering with M. fusca and M. perspicillata in order. Phylogenetic analysis suggests that Mareca falcata and M. strepera are sister groups, differing from previous studies. Results firstly indicate that Clangula hyemalis and Somateria mollissima are sister groups, suggesting a potentially skewed phylogenetic relationship may have been overlooked in earlier analyses relying solely on mitochondrial genomes. Our results provide new mitogenome data to support further phylogenetic and taxonomic studies of Anseriformes. [ABSTRACT FROM AUTHOR]

PSA screening has led to an over-diagnosis of prostate cancer (PCa) and unnecessary biopsies of benign conditions due to its low cancer specificity. Consequently, more accurate, preferentially non-invasive, tests are needed. We aim to evaluate the potential of semen sEV (small extracellular vesicles) tsRNAs (tRNA-derived small RNAs) as PCa indicators. Initially, following a literature review in the OncotRF database and high-throughput small RNA-sequencing studies in PCa tissue together with the sncRNA profile in semen sEVs, we selected four candidate 5′tRF tsRNAs for validation as PCa biomarkers. RT-qPCR analysis in semen sEVs from men with moderately elevated serum PSA levels successfully shows that the differential expression of the four tRFs between PCa and healthy control groups can be detected in a non-invasive manner. The combined model incorporating PSA and specific tRFs (5′-tRNA-Glu-TTC-9-1_L30 and 5′-tRNA-Val-CAC-3-1_L30) achieved high predictive accuracy in identifying samples with a Gleason score ≥ 7 and staging disease beyond IIA, supporting that the 5′tRF fingerprint in semen sEV can improve the PSA predictive value to discriminate between malignant and indolent prostate conditions. The in silico study allowed us to map target genes for the four 5′tRFs possibly involved in PCa. Our findings highlight the synergistic use of multiple biomarkers as an efficient approach to improve PCa screening and prognosis. [ABSTRACT FROM AUTHOR]

Intestinal tuft cells are critical for anti-helminth parasite immunity because they produce IL-25, which triggers IL-13 secretion by activated group 2 innate lymphoid cells (ILC2s) to expand both goblet and tuft cells. We show that epithelial Elp3, a tRNA-modifying enzyme, promotes tuft cell differentiation and is consequently critical for IL-25 production, ILC2 activation, goblet cell expansion and control of Nippostrongylus brasiliensis helminth infection in mice. Elp3 is essential for the generation of intestinal immature tuft cells and for the IL-13-dependent induction of glycolytic enzymes such as Hexokinase 1 and Aldolase A. Importantly, loss of epithelial Elp3 in the intestine blocks the codon-dependent translation of the Gator1 subunit Nprl2, an mTORC1 inhibitor, which consequently enhances mTORC1 activation and stabilizes Atf4 in progenitor cells. Likewise, Atf4 overexpression in mouse intestinal epithelium blocks tuft cell differentiation in response to intestinal helminth infection. Collectively, our data define Atf4 as a negative regulator of tuft cells and provide insights into promotion of intestinal type 2 immune response to parasites through tRNA modifications. Synopsis: Epithelial host response mechanisms against parasites remain poorly understood. This work identifies the tRNA-modifying enzyme Elp3 as a critical promoter of intestinal tuft cell expansion and type 2 immunity in mice, mounting protection against helminth infection. Elp3 is required for the anti-helminth immune response in the intestine. Epithelial Elp3 depletion inhibits tuft cell differentiation. Elp3 deficiency decreases mRNA translation of mTORC1 inhibitor Nprl2, potentiating mTORC1-Atf4 signaling. Atf4 overexpression in the intestinal epithelium results in incomplete response to helminth infections due to defective tuft cell amplification. tRNA-modifying enzyme Elp3 is required for protective type 2 immunity in the intestinal epithelium upon helminth infection, supporting tuft cell expansion and goblet cell function. [ABSTRACT FROM AUTHOR]

Background: The Bucephalidae is a large family of digenean trematodes but most previous analyses of its phylogenetic position have relied on a single mitochondrial gene or morphological features. Mitochondrial genomes (mitogenomes) remain unavailable for the entire family. To address this, we sequenced the complete mitogenome of Dollfustrema vaneyi and analyzed the phylogenetic relationships with other trematodes. Results: The circular genome of Dollfustrema vaneyi spanned 14,959 bp and contained 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and a major non-coding region. We used concatenated amino acid and nucleotide sequences of all 36 genes for phylogenetic analyses, conducted using MrBayes, IQ-TREE and PhyloBayes. We identified pronounced topological instability across different analyses. The addition of recently sequenced two mitogenomes for the Aspidogastrea subclass along with the use of a site-heterogeneous model stabilized the topology, particularly the positions of Azygiidae and Bucephalidae. The stabilized results indicated that Azygiidae was the closest lineage to Bucephalidae in the available dataset, and together, they clustered at the base of the Plagiorchiida. Conclusions: Our study provides the first comprehensive description and annotation of the mitochondrial genome for the Bucephalidae family. The results indicate a close phylogenetic relationship between Azygiidae and Bucephalidae, and reveal their basal placement within the order Plagiorchiida. Furthermore, the inclusion of Aspidogastrea mitogenomes and the site-heterogeneous model significantly improved the topological stability. These data will provide key molecular resources for future taxonomic and phylogenetic studies of the family Bucephalidae. [ABSTRACT FROM AUTHOR]

The ribosome utilizes hydrogen bonding between mRNA codons and aminoacyl-tRNAs to ensure rapid and accurate protein production. Chemical modification of mRNA nucleobases can adjust the strength and pattern of this hydrogen bonding to alter protein synthesis. We investigate how the N1-methylpseudouridine (m1Ψ) modification, commonly incorporated into therapeutic and vaccine mRNA sequences, influences the speed and fidelity of translation. We find that m1Ψ does not substantially change the rate constants for amino acid addition by cognate tRNAs or termination by release factors. However, we also find that m1Ψ can subtly modulate the fidelity of amino acid incorporation in a codon-position and tRNA dependent manner in vitro and in human cells. Our computational modeling shows that altered energetics of mRNA:tRNA interactions largely account for the context dependence of the low levels of miscoding we observe on Ψ and m1Ψ containing codons. The outcome of translation on modified mRNA bases is thus governed by the sequence context in which they occur. Chemical modification of mRNA nucleobases alters hydrogen bonding during translation. Here the authors show that the N1-methylpseudouridine (m1ψ), used in therapeutics, does not change translation rate but modestly modulates its fidelity in a codon-position and tRNA dependent manner. [ABSTRACT FROM AUTHOR]

The mitochondrial genome is recognized for its utility in insect molecular research, due to its distinctive features, including fast evolutionary rate, low recombination, and maternal inheritance. In this study, we explored the mitochondrial genome in the context of the leafhopper subfamily Evacanthinae, which is significant in the agricultural pest sector due to the direct and indirect damage caused to plants by its species. We present complete mitochondrial genome sequences for three species: Evacanthus bivittatus, Carinata ganga, and Carinata recurvata. Comparative analyses of nucleotide composition, codon usage of protein coding genes (PCGs), nucleotide diversity, transfer RNA secondary structure, and gene overlap were conducted. To understand phylogenetic relationships among species, we constructed phylogenetic trees using nucleotide sequences from the 13 PCGs of the genomes newly sequenced in this study alongside other available leafhopper genome sequences. Phylogenetic analysis supported monophyly of the Evacanthinae subfamily and suggested a close relationship between the Evacanthini and Nirvanini tribes. Our research indicates that anticodon differences in trnSl are insufficient to serve as taxonomic markers for distinguishing between these two tribes. This study contributes valuable genetic data supporting ongoing investigations into genetic diversity, molecular evolution, and species identification, while laying groundwork for future taxonomic and evolutionary endeavors within Cicadellidae. [ABSTRACT FROM AUTHOR]

tRNAs are codon decoders that convert the transcriptome into the proteome. The field of tRNA research is excited by the increasing discovery of specific tRNA modifications that are installed at specific, evolutionarily conserved positions by a set of specialized tRNA-modifying enzymes and the biogenesis of tRNA-derived regulatory fragments (tsRNAs) which exhibit copious activities through multiple mechanisms. Dysregulation of tRNA modification usually has pathological consequences, a phenomenon referred to as "tRNA modopathy". Current evidence suggests that certain tRNA-modifying enzymes and tsRNAs may serve as promising diagnostic biomarkers and therapeutic targets, particularly for chemoresistant cancers. In this review, we discuss the latest discoveries that elucidate the molecular mechanisms underlying the functions of clinically relevant tRNA modifications and tsRNAs, with a focus on malignancies. We also discuss the therapeutic potential of tRNA/tsRNA-based therapies, aiming to provide insights for the development of innovative therapeutic strategies. Further efforts to unravel the complexities inherent in tRNA biology hold the promise of yielding better biomarkers for the diagnosis and prognosis of diseases, thereby advancing the development of precision medicine for health improvement. [ABSTRACT FROM AUTHOR]

Modern life is essentially homochiral, containing D-sugars in nucleic acid backbones and L-amino acids in proteins. Since coded proteins are theorized to have developed from a prebiotic RNA World, the homochirality of L-amino acids observed in all known life presumably resulted from chiral transfer from a homochiral D-RNA World. This transfer would have been mediated by aminoacyl-RNAs defining the genetic code. Previous work on aminoacyl transfer using tRNA mimics has suggested that aminoacylation using D-RNA may be inherently biased toward reactivity with L-amino acids, implying a deterministic path from a D-RNA World to L-proteins. Using a model system of self-aminoacylating D-ribozymes and epimerizable activated amino acid analogs, we test the chiral selectivity of 15 ribozymes derived from an exhaustive search of sequence space. All of the ribozymes exhibit detectable selectivity, and a substantial fraction react preferentially to produce the D-enantiomer of the product. Furthermore, chiral preference is conserved within sequence families. These results are consistent with the transfer of chiral information from RNA to proteins but do not support an intrinsic bias of D-RNA for L-amino acids. Different aminoacylation structures result in different directions of chiral selectivity, such that L-proteins need not emerge from a D-RNA World. Life is homochiral, and prior work posited that D-RNA may interact preferentially with L-amino acids. Here, the authors show that self-aminoacylating D-RNAs do not have an intrinsic preference for L-amino acids, suggesting that either handedness could emerge from early living systems. [ABSTRACT FROM AUTHOR]

More than 100 RNA chemical modifications to cellular RNA have been identified. N6-methyladenosine (m6A) is the most prevalent modification of mRNA. RNA modifications have recently attracted significant attention due to their critical role in regulating mRNA processing and metabolism. tRNA and rRNA rank among the most heavily modified RNAs, and their modifications are essential for maintaining their structure and function. With our advanced understanding of RNA modifications, increasing evidence suggests RNA modifications are important in regulating various aspects of insect life. In this review, we will summarize recent studies investigating the impact of RNA modifications in insects, particularly highlighting the role of m6A in insect development, reproduction, and adaptation to the environment. [ABSTRACT FROM AUTHOR]

The aminoacyl‐tRNA synthetases (aaRS) are a large group of enzymes that implement the genetic code in all known biological systems. They attach amino acids to their cognate tRNAs, moonlight in various translational and non‐translational activities beyond aminoacylation, and are linked to many genetic disorders. The aaRS have a subtle ontology characterized by structural and functional idiosyncrasies that vary from organism to organism, and protein to protein. Across the tree of life, the 22 coded amino acids are handled by 16 evolutionary families of Class I aaRS and 21 families of Class II aaRS. We introduce AARS Online, an interactive Wikipedia‐like tool curated by an international consortium of field experts. This platform systematizes existing knowledge about the aaRS by showcasing a taxonomically diverse selection of aaRS sequences and structures. Through its graphical user interface, AARS Online facilitates a seamless exploration between protein sequence and structure, providing a friendly introduction to the material for non‐experts and a useful resource for experts. Curated multiple sequence alignments can be extracted for downstream analyses. Accessible at www.aars.online, AARS Online is a free resource to delve into the world of the aaRS. [ABSTRACT FROM AUTHOR]

Pseudouridine (Ψ) is an ubiquitous RNA modification, present in the tRNAs and rRNAs of species across all domains of life. Conserved pseudouridine synthases modify the mRNAs of diverse eukaryotes, but the modification has yet to be identified in bacterial mRNAs. Here, we report the discovery of pseudouridines in mRNA from E. coli. By testing the mRNA modification capacity of all 11 known pseudouridine synthases, we identify RluA as the predominant mRNA-modifying enzyme. RluA, a known tRNA and 23S rRNA pseudouridine synthase, modifies at least 31 of the 44 high-confidence sites we identified in E. coli mRNAs. Using RNA structure probing data to inform secondary structures, we show that the target sites of RluA occur in a common sequence and structural motif comprised of a ΨURAA sequence located in the loop of a short hairpin. This recognition element is shared with previously identified target sites of RluA in tRNAs and rRNA. Overall, our work identifies pseudouridine in key mRNAs and suggests the capacity of Ψ to regulate the transcripts that contain it. Author summary: While RNAs are initially transcribed using only the nucleotides A, G, C and U, these nucleotides can be enzymatically modified into many different post-transcriptional modifications. tRNAs and rRNAs across all domains of life are modified with pseudouridine, an isoform of uridine that is inserted by highly conserved enzymes. In many eukaryotes, it has been demonstrated that some of these enzymes can also insert pseudouridines in mRNA, where they are poised to impact gene expression through their effects on the transcript. Here we show that protein-coding transcripts in E. coli are also modified with pseudouridine, indicating that mRNA pseudouridylation is also a highly conserved activity. RluA is the main mRNA pseudouridine synthase, introducing the modification into the transcripts of dozens of protein coding genes with high specificity. Its target sites are defined by a combined sequence and secondary structure motif. Two additional enzymes, RluC and RluD, introduce a few additional sites. All three of these enzymes belong to the same sub-family of pseudouridine synthases, and homologs of these also modify mRNAs in humans. Thus, mRNA modification by these enzymes might be a conserved activity with the capacity to impact gene regulation. [ABSTRACT FROM AUTHOR]

Lycophytes and ferns represent one of the earliest-diverging lineages of vascular plants, with the Lycopodiaceae family constituting the basal clade among lycophytes. In this research, we successfully assembled and annotated the complete Lycopodium japonicum Thunb. (L. japonicum) mitochondrial genome (mitogenome) utilizing PacBio HiFi sequencing data, resulting in a single circular molecule with a size of 454,458 bp. 64 unique genes were annotated altogether, including 34 protein-coding genes, 27 tRNAs and 3 rRNAs. It also contains 32 group II introns, all of which undergo cis-splicing. We identified 195 simple sequence repeats, 1,948 dispersed repeats, and 92 tandem repeats in the L. japonicum mitogenome. Collinear analysis indicated that the mitogenomes of Lycopodiaceae are remarkably conserved compared to those of other vascular plants. We totally identified 326 RNA editing sites in 31 unique protein-coding genes with 299 sites converting cytosine to uracil and 27 sites the reverse. Notably, the L. japonicum mitogenome has small amounts foreign DNA from plastid or nuclear origin, accounting for only 2.81% of the mitogenome. The maximum likelihood phylogenetic analysis based on 23 diverse land plant mitogenomes and plastid genomes supports the basal position of lycophytes within vascular plants and they form a sister clade to all other vascular lineages, which is consistent with the PPG I classification system. As the first reported mitogenome of Lycopodioideae subfamily, this study enriches our understanding of Lycopodium mitogenomes, and sets the stage for future research on mitochondrial diversity and evolution within the lycophytes and ferns. [ABSTRACT FROM AUTHOR]

Transfer RNAs (tRNA) are essential small non-coding RNAs that enable the translation of genomic information into proteins in all life forms. The principal function of tRNAs is to bring amino acid building blocks to the ribosomes for protein synthesis. In the ribosome, tRNAs interact with messenger RNA (mRNA) to mediate the incorporation of amino acids into a growing polypeptide chain following the rules of the genetic code. Accurate interpretation of the genetic code requires tRNAs to carry amino acids matching their anticodon identity and decode the correct codon on mRNAs. Errors in these steps cause the translation of codons with the wrong amino acids (mistranslation), compromising the accurate flow of information from DNA to proteins. Accumulation of mutant proteins due to mistranslation jeopardizes proteostasis and cellular viability. However, the concept of mistranslation is evolving, with increasing evidence indicating that mistranslation can be used as a mechanism for survival and acclimatization to environmental conditions. In this review, we discuss the central role of tRNAs in modulating translational fidelity through their dynamic and complex interplay with translation factors. We summarize recent discoveries of mistranslating tRNAs and describe the underlying molecular mechanisms and the specific conditions and environments that enable and promote mistranslation. [ABSTRACT FROM AUTHOR]

Exportin5 (Exp5) is the major miRNA nuclear export factor and recognizes structural features of pre-miRNA hairpins, while it also exports other minihelix-containing RNAs. In Drosophila, Exp5 is suggested to play a major role in tRNA export because the gene encoding the canonical tRNA export factor Exportin-t is missing in its genome. To understand molecular functions of fly Exp5, we studied the Exp5/RNA interactome in the cell line S2R + using the crosslinking and immunoprecipitation (CLIP) technology. The CLIP experiment captured known substrates such as tRNAs and miRNAs and detected candidates of novel Exp5 substrates including various mRNAs and long non-coding RNAs (lncRNAs). Some mRNAs and lncRNAs enriched PAR-CLIP tags compared to their expression levels, suggesting selective binding of Exp5 to them. Intronless mRNAs tended to enrich PAR-CLIP tags; therefore, we proposed that Exp5 might play a role in the export of specific classes of mRNAs/lncRNAs. This result suggested that Drosophila Exp5 might have a wider variety of substrates than initially thought. Surprisingly, Exp5 CLIP reads often contained sequences corresponding to the flanking 50 -leaders and 30 -trailers of tRNAs, which were thought to be removed prior to nuclear export. In fact, we found pre-tRNAs before endprocessing were present in the cytoplasm, supporting the idea that tRNA end-processing is a cytoplasmic event. In summary, our results provide a genome-wide list of Exp5 substrate candidates and suggest that flies may lack a mechanism to distinguish pre-tRNAs with or without the flanking sequences. [ABSTRACT FROM AUTHOR]

Leber’s Hereditary Optic Neuropathy (LHON) is a rare, maternally inherited eye disease, predominantly due to the degeneration of retinal ganglion cells (RGCs). It is associated with a mitochondrial DNA (mtDNA) point mutation. Our previous study identified that the m.15927G > A homoplasmic mutation damaged the highly conserved base pairing (28C42G) in anticodon stem of tRNAThr, caused deficient t6 A modification and significantly decreased efficiency in aminoacylation and steady-state levels of tRNAThr, and led to mitochondrial dysfunction. Meanwhile, mechanisms underlying mtDNA mutations regulate intracellular signaling related to mitochondrial and cellular integrity are less explored. Here, we manifested that defective nucleotide modification induced by the m.15927G > A mutation interfered with the expression of nuclear genes involved in cytoplasmic proteins essential for oxidative phosphorylation system (OXPHOS), thereby impacting the assemble and integrity of OXPHOS complexes. As a result of these mitochondrial dysfunctions, there was an imbalance in mitochondrial dynamics, particularly distinguished by an increased occurrence of mitochondrial fission. Excessive fission compromised the autophagy process, including the initiation phase, formation, and maturation of autophagosomes. Both Parkin-mediated mitophagy and receptor-dependent mitophagy were significantly impaired in cybrids haboring the m.15927G > A mutation. These changes facilitated intrinsic apoptosis, as indicated by increased cytochrome c release and elevated levels of apoptosis-associated proteins (e.g., BAK, BAX, cleaved caspase 9, cleaved caspase 3, and cleaved PARP) in the mutant cybrids. This study demonstrates that the m.15927G > A mutation contributes to LHON by dysregulating OXPHOS biogenesis, aberrant quality control, increased autophagy, inhibited mitophagy, and abnormal apoptosis. [ABSTRACT FROM AUTHOR]

Simple Summary: A total of 37 mitochondrial genes appears in a certain arrangement in most insects, and the transcription of this conserved mitochondrial genome (mitogenome) has been well-studied in species like Drosophila melanogaster, Erthesina fullo and Coridius chinensis. However, transcription of the mitogenome with extensive gene rearrangement has rarely been studied. In this research, we sequenced the mitogenome and mitochondrial transcriptome of Sericothrips houjii (Thysanoptera: Thripidae: Sericothripinae). The mitogenome of S. houjii exhibited extensive gene rearrangement and contained two control regions (CRs) with sequence repeats. Compared to the insect mitogenome with a typical gene order, the mitogenomic feature, relative gene expression level, transcriptional model and post-transcriptional cleavage of S. houjii were quite different. Unlike other insects where ribosomal RNA (rRNA) is typically highly expressed, ND4/ND4L in S. houjii exhibits the highest expression. Both strands of this mitogenome were entirely transcribed and the bicistronic messenger RNA (mRNA) COI/ND3 was reported for the first time in insects. Our study provides new insights into the transcriptional and post-transcriptional regulation processes in the insect mitogenome with extensive gene rearrangement and duplicated CRs. The mitochondrial genome (mitogenome) of Thysanoptera has extensive gene rearrangement, and some species have repeatable control regions. To investigate the characteristics of the gene expression, transcription and post-transcriptional processes in such extensively gene-rearranged mitogenomes, we sequenced the mitogenome and mitochondrial transcriptome of Sericothrips houjii to analyze. The mitogenome was 14,965 bp in length and included two CRs contains 140 bp repeats between COIII-trnN (CR1) and trnT-trnP (CR2). Unlike the putative ancestral arrangement of insects, S. houjii exhibited only six conserved gene blocks encompassing 14 genes (trnL2-COII, trnD-trnK, ND2-trnW, ATP8-ATP6, ND5-trnH-ND4-ND4L and trnV-lrRNA). A quantitative transcription map showed the gene with the highest relative expression in the mitogenome was ND4-ND4L. Based on analyses of polycistronic transcripts, non-coding RNAs (ncRNAs) and antisense transcripts, we proposed a transcriptional model of this mitogenome. Both CRs contained the transcription initiation sites (TISs) and transcription termination sites (TTSs) of both strands, and an additional TIS for the majority strand (J-strand) was found within antisense lrRNA. The post-transcriptional cleavage processes followed the "tRNA punctuation" model. After the cleavage of transfer RNAs (tRNAs), COI and ND3 matured as bicistronic mRNA COI/ND3 due to the translocation of intervening tRNAs, and the 3′ untranslated region (UTR) remained in the mRNAs for COII, COIII, CYTB and ND5. Additionally, isoform RNAs of ND2, srRNA and lrRNA were identified. In summary, the relative mitochondrial gene expression levels, transcriptional model and post-transcriptional cleavage process of S. houjii are notably different from those insects with typical mitochondrial gene arrangements. In addition, the phylogenetic tree of Thripidae including S. houjii was reconstructed. Our study provides insights into the phylogenetic status of Sericothripinae and the transcriptional and post-transcriptional regulation processes of extensively gene-rearranged insect mitogenomes. [ABSTRACT FROM AUTHOR]