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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]

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]

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]

The yellowtail snapper Ocyurus chrysurus Bloch, 1791 (Perciformes: Lutjanidae) is targeted by recreational or commercial fisheries across much of its range in the western Atlantic Ocean, from Massachusetts, USA, to São Paulo, Brazil. Genomic resources to support conservation of this species are needed. Here, we assembled, annotated and characterised in detail the mitochondrial genome of O. chrysurus. The mitochondrial genome of O. chrysurus is 16,502 bp in length and encodes 13 protein-coding genes (PCGs), 22 transfer RNA genes (tRNA), and two ribosomal RNA genes (rRNA). The putative control region, 827 bp in length, is located between the tRNA P and tRNA F genes. Most genes are encoded on the positive strand, while a single PCG (nad6) and six tRNA genes (in order from 5′ to 3′: tRNA N, tRNA P, tRNA E, tRNA Q, tRNA C and tRNA Y) are encoded on the negative strand. All tRNA have a typical 'cloverleaf' secondary structure except tRNA S1 (Serine1) that lacks the D arm. Calculated Ka/Ks values of all PCGs were <1, suggesting that these genes are under purifying selection. A maximum likelihood (ML) phylogenetic analysis confirmed monophyly of the family Lutjanidae and indicated that the genus Lutjanus is paraphyletic given that O. chrysurus clustered together with Lutjanus peru and L. guttatus in a strongly supported clade. Data published herein will inform conservation and management strategies for this fished snapper. [ABSTRACT FROM AUTHOR]

Among isopods (order Isopoda), species belonging to the suborder Oniscidea are the most successful colonisers of terrestrial habitats and the largest family in this order is Armadillidae. In this study, the complete mitochondrial genome of an armadillid isopod, Cubaris murina, was assembled and characterised in detail for the first time. The gene order and secondary structure of transfer RNA (tRNA) genes were compared among isopods. The 14,212-bp-long mitogenome consists of 13 protein-coding genes, 20 tRNAs, two ribosomal RNAs, and four non-coding regions (NCRs), each longer than 100 bp. Two tRNA genes (trnG and trnI) were missing and several other tRNA genes exhibited secondary structures that departed from the typical cloverleaf pattern. Deletions and nucleotide insertions in the T- and D-arms and mismatches in the acceptor stem were commonly observed in these tRNAs. Notably, the mitogenome of C. murina exhibited a new gene order containing four tRNA rearrangements, and differences in the location of the NCRs were detected when compared to those reported for other isopods, including oniscid species. Interestingly, the second NCR had a similar number of secondary structures and repetitive elements to those reported in the control region of other crustaceans. The novel gene order, tRNA secondary structures, and NCR organisation reported herein for the mitogenome of C. murina confirms the hyper-variability of mitochondrial genomes in isopods. This study is the first to characterise in detail the mitogenome of the invasive isopod C. murina, belonging to the species-rich family Armadillidae within the Isopoda, and advances our understanding of mitochondrial genome architecture in isopods. [ABSTRACT FROM AUTHOR]

Post‐transcriptional tRNA modifications contribute to the decoding efficiency of tRNAs by supporting codon recognition and tRNA stability. Recent work shows that the molecular and cellular functions of tRNA modifications and tRNA‐modifying‐enzymes are linked to brain development and neurological disorders. Lack of these modifications affects codon recognition and decoding rate, promoting protein aggregation and translational stress response pathways with toxic consequences to the cell. In this review, we discuss the peculiarity of local translation in neurons, suggesting a role for fine‐tuning of translation performed by tRNA modifications. We provide several examples of tRNA modifications involved in physiology and pathology of the nervous system, highlighting their effects on protein translation and discussing underlying mechanisms, like the unfolded protein response (UPR), ribosome quality control (RQC), and no‐go mRNA decay (NGD), which could affect neuronal functions. We aim to deepen the understanding of the roles of tRNA modifications and the coordination of these modifications with the protein translation machinery in the nervous system. [ABSTRACT FROM AUTHOR]

Transfer RNA-derived small RNAs (tsRNAs), a recently identified subclass of small non-coding RNAs (sncRNAs), emerge through the cleavage of mature transfer RNA (tRNA) or tRNA precursors mediated by specific enzymes. The tumor necrosis factor (TNF) protein, a signaling molecule produced by activated macrophages, plays a pivotal role in systemic inflammation. Its multifaceted functions include the capacity to eliminate or hinder tumor cells, enhance the phagocytic capabilities of neutrophils, confer resistance against infections, induce fever, and prompt the production of acute phase proteins. Notably, four TNF-related tsRNAs have been conclusively linked to distinct diseases. Examples include 5′tiRNA-Gly in skeletal muscle injury, tsRNA-21109 in systemic lupus erythematosus (SLE), tRF-Leu-AAG-001 in endometriosis (EMs), and tsRNA-04002 in intervertebral disk degeneration (IDD). These tsRNAs exhibit the ability to suppress the expression of TNF-α. Additionally, KEGG analysis has identified seven tsRNAs potentially involved in modulating the TNF pathway, exerting their influence across a spectrum of non-cancerous diseases. Noteworthy instances include aberrant tiRNA-Ser-TGA-001 and tRF-Val-AAC-034 in intrauterine growth restriction (IUGR), irregular tRF-Ala-AGC-052 and tRF-Ala-TGC-027 in obesity, and deviant tiRNA-His-GTG-001, tRF-Ser-GCT-113, and tRF-Gln-TTG-035 in irritable bowel syndrome with diarrhea (IBS-D). This comprehensive review explores the biological functions and mechanisms of tsRNAs associated with the TNF signaling pathway in both cancer and other diseases, offering novel insights for future translational medical research. [ABSTRACT FROM AUTHOR]

• arg-tRNAarg is essential for protein synthesis in the ribosome. • arg-tRNAarg is also used for post-translational modification of proteins in all eukaryotes. • Stable arg-tRNA are very interesting tools to better understand the specificity of the arginyl transferases enzyme. • The use of chemical strategies to synthetize stable analogs allows to obtain the two regioisomers of aminoacyl-tRNA. In this report, non-isomerisable analogs of arginine tRNA (Arg-triazole-tRNA) have been synthesized as tools to study tRNA-dependent aminoacyl-transferases. The synthesis involves the incorporation of 1,4 substituted-1,2,3 triazole ring to mimic the ester bond that connects the amino acid to the terminal adenosine in the natural substrate. The synthetic procedure includes (i) a coupling between 2′- or 3′-azido-adenosine derivatives and a cytidine phosphoramidite to access dinucleotide molecules, (ii) Cu-catalyzed cycloaddition reactions between 2′- or 3′-azido dinucleotide in the presence of an alkyne molecule mimicking the arginine, providing the corresponding Arg-triazole-dinucleotides, (iii) enzymatic phosphorylation of the 5′-end extremity of the Arg-triazole-dinucleotides with a polynucleotide kinase, and (iv) enzymatic ligation of the 5′-phosphorylated dinucleotides with a 23-nt RNA micro helix that mimics the acceptor arm of arg-tRNA or with a full tRNAarg. Characterization of nucleoside and nucleotide compounds involved MS spectrometry, 1H, 13C and 31P NMR analysis. This strategy allows to obtain the pair of the two stable regioisomers of arg-tRNA analogs (2′ and 3′) which are instrumental to explore the regiospecificity of arginyl transferases enzyme. In our study, a first binding assay of the arg-tRNA micro helix with the Arginyl-tRNA-protein transferase 1 (ATE1) was performed by gel shift assays. [ABSTRACT FROM AUTHOR]

Comparative mitogenomics and its evolutionary relationships within Bryconidae remains largely unexplored. To bridge this gap, this study assembled 15 mitogenomes from 11 Bryconidae species, including five newly sequenced. Salminus mitogenomes, exceeding 17,700 bp, exhibited the largest size, contrasting with a median size of 16,848 bp in the remaining species (Brycon and Chilobrycon). These mitogenomes encode 37 typical mitochondrial genes, including 13 protein-coding, 2 ribosomal RNA, and 22 transfer RNA genes, and exhibit the conserved gene arrangement found in most fish species. Phylogenetic relationships, based on the maximum-likelihood method, revealed that the trans-Andean species (found in northwestern South America) clustered into two main sister clades. One clade comprised the trans-Andean species from the Pacific slope, Brycon chagrensis and Chilobrycon deuterodon. The other clade grouped the trans-Andean species from the Magdalena-Cauca Basin Brycon moorei and Salminus affinis, with their respective cis-Andean congeners (found in eastern South America), with Brycon rubricauda as its sister clade. Since the current members of Brycon are split in three separated lineages, the systematic classification of Bryconidae requires further examination. This study provides novel insights into mitogenome characteristics and evolutionary pathways within Bryconidae, standing as crucial information for prospective phylogenetic and taxonomic studies, molecular ecology, and provides a valuable resource for environmental DNA applications. [ABSTRACT FROM AUTHOR]

Ophiuroids, the most diverse group of echinoderms, inhabit a vast array of ecological niches and play vital roles in benthic ecosystems as suspension feeders, scavengers and opportunists. Despite the important ecological roles played by Ophiuroidea, their evolutionary history and phylogenetic relationship is not yet fully understood. In this study, 47 mitochondrial genomes of ophiuroids, including 21 newly sequenced ones, were analyzed. tRNA duplication was firstly discovered in four species and a new start codon was identified for Ophiuroidea. Eighteen phylogenetic trees based on mitochondrial genomes consistently supported two major lineages, Ophintegrida and Euryophiurida. It further confirmed the monophyly of Euryalida and Ophiurida, respectively, as well as families represented by multiple species. Among 18 trees, only the two ML trees based on amino acid sequences using IQtree method supported monophyly of Amphilepidida and Ophiacanthida, consistent with current phylogenetic system of Ophiuroidea. This result highlighted the effect of phylogenetic analysis methods and datasets on tree topology, indicating that amino acid sequence data maybe more suitable for higher taxonomic level phylogenetic analysis of ophiuroids than nucleotide sequences. Four new gene orders of 13 proteincoding genes + two rRNAs and 12 new gene orders of all 37 genes were identified. Mitochondrial gene orders were highly variable in Ophiacanthida, but were extremely conserved in Eurylida. Additionally, both branch lengths and estimated positive selection varied among the four orders, and a positive relationship between branch lengths and mitochondrial gene rearrangement rates was revealed, suggesting distinctly different evolutionary history among the four major clades of Ophiuroidea. Overall, we (1) reconstructed the phylogenetic relationship based on mitochondrial genome, supporting the current phylogenetic system in Ophiuroidea, (2) revealed a high variability in mitochondrial gene rearrangement among the four orders, (3) provided the first evidence to link gene rearrangement and nucleotide substitution in Echinodermata. [ABSTRACT FROM AUTHOR]

Structural variants play an important role in evolutionary processes. Besides, they constitute a large source of inter individual genetic variation that might represent a major factor in the aetiology of complex, multifactorial traits. Their importance in adaptation is becoming increasingly evident in literature. Yet, the characterization of the genomic landscape of structural variants in local breeds remains scarce to date. Herein, we investigate patterns and gene annotation of structural variants in the Creole cattle from Guadeloupe breed using whole genome sequences from 23 bulls representative of the population. In total, we detected 32821 ascertained SV defining 15258 regions, representing ~ 17% of the Creole cattle genome. Among these, 6639 regions have not been previously reported in the Database of Genomic Variants archive. Average number of structural variants detected per individual in the studied population is in the same order of magnitude of that observed in indicine populations and higher than that reported in taurine breeds. We observe an important within-individual variability where approximately half of the detected structural variants have low frequency (MAF < 0.25). Most of the detected structural variants (55%) occurred in intergenic regions. Genic structural variants overlapped with 7793 genes and the predicted effect of most of them is ranked as "modifier". Among the structural variants that were predicted to have a high functional impact on the protein, a 5.5 Kb in length, highly frequent deletion on chromosome 2, affects ALPI, a gene associated with the interaction between gut microbiota and host immune system. The 6639 newly identified structural variants regions include three deletions and three duplications shared by more than 80% of individuals that are significantly enriched for genes related to tRNA threonylcarbamoyladenosine metabolic process, important for temperature adaptation in thermophilic organisms, therefore suggesting a potential role in the thermotolerance of Creole cattle from Guadeloupe cattle to tropical climate. Overall, highly frequent structural variants that are specific to the Creole cattle population encompass olfactory receptor and immunity genes as well as genes involved in muscle tone, muscle development and contraction. Beyond mapping and characterizing structural variants in the Creole cattle from Guadeloupe breed, this study provides valuable information for a better understanding of the potential role of chromosomal rearrangements in adaptive traits in cattle. [ABSTRACT FROM AUTHOR]

The IUCN Red List of Threatened Species contains 175 Brazilian bat species that are threatened by extinction in some degree. From this perspective, it is essential to expand the knowledge about the genetic diversity of vulnerable bats. Genomic sequencing can be useful to generate robust and informative genetic references, increasing resolution when analyzing relationships among populations, species, or higher taxonomic levels. In this study, we sequenced and characterized in detail the first complete mitochondrial genomes of Furipterus horrens, Lonchorhina aurita, and Natalus macrourus, and investigated their phylogenetic position based on amino acid sequences of protein-coding genes (PCGs). The mitogenomes of these species are 16,516, 16,697, and 16,668 bp in length, respectively, and each comprises 13 PCGs, 22 tRNA genes, two rRNA genes, and a putative control region (CR). In the three species, genes were arranged similarly to all other previously described bat mitogenomes, and nucleotide composition was also consistent with the reported range. The length and arrangement of rrnS and rrnL were also consistent with those of other bat species, showing a positive AT-skew and a negative GC-skew. Except for trnS1, for which we did not observe the DHU arm, all other tRNAs showed the cloverleaf secondary structure in the three species. In addition, the mitogenomes showed minor differences in start and stop codons, and in all PCGs, codons ending in adenine were more common compared to those ending in guanine. We found that PCGs of the three species use multiple codons to encode each amino acid, following the previously documented pattern. Furthermore, all PCGs are under purifying selection, with atp8 experiencing the most relaxed purifying selection. Considering the phylogenetic reconstruction, F. horrens was recovered as sister to Noctilio leporinus, L. aurita and Tonatia bidens shared a node within Phyllostomidae, and N. macrourus appeared as sister to Molossidae and Vespertilionidae. [ABSTRACT FROM AUTHOR]

Emerging evidence indicates that transfer RNA (tRNA)‐derived small RNAs (tsRNAs), originated from tRNA with high abundance RNA modifications, play an important role in many complex physiological and pathological processes. However, the biological functions and regulatory mechanisms of modified tsRNAs in cancer remain poorly understood. Here, it is screened for and confirmed the presence of a novel m7G‐modified tsRNA, m7G‐3′‐tiRNA LysTTT (mtiRL), in a variety of chemical carcinogenesis models by combining small RNA sequencing with an m7G small RNA‐modified chip. Moreover, it is found that mtiRL, catalyzed by the tRNA m7G‐modifying enzyme mettl1, promotes bladder cancer (BC) malignancy in vitro and in vivo. Mechanistically, mtiRL is found to specifically bind the oncoprotein Annexin A2 (ANXA2) to promote its Tyr24 phosphorylation by enhancing the interactions between ANXA2 and Yes proto‐oncogene 1 (Yes1), leading to ANXA2 activation and increased p‐ANXA2‐Y24 nuclear localization in BC cells. Together, these findings define a critical role for mtiRL and suggest that targeting this novel m7G‐modified tsRNA can be an efficient way for to treat BC. [ABSTRACT FROM AUTHOR]

We leverage machine learning approaches to adapt nanopore sequencing basecallers for nucleotide modification detection. We first apply the incremental learning (IL) technique to improve the basecalling of modification-rich sequences, which are usually of high biological interest. With sequence backbones resolved, we further run anomaly detection (AD) on individual nucleotides to determine their modification status. By this means, our pipeline promises the single-molecule, single-nucleotide, and sequence context-free detection of modifications. We benchmark the pipeline using control oligos, further apply it in the basecalling of densely-modified yeast tRNAs and E.coli genomic DNAs, the cross-species detection of N6-methyladenosine (m6A) in mammalian mRNAs, and the simultaneous detection of N1-methyladenosine (m1A) and m6A in human mRNAs. Our IL-AD workflow is available at: https://github.com/wangziyuan66/IL-AD. Here the authors adapt nanopore sequencing basecallers to detect RNA modifications. The authors first apply incremental learning to resolve modification-disturbed basecalling, then use anomaly detection to assess nucleotide modification status. [ABSTRACT FROM AUTHOR]

Despite the highly conserved nature of the genetic code, the frequency of usage of each codon can vary significantly. The evolution of codon usage is shaped by two main evolutionary forces: mutational bias and selection pressures. These pressures can be driven by environmental factors, but also by the need for efficient translation, which depends heavily on the concentration of transfer RNAs (tRNAs) within the cell. The data presented here supports the proposal that tRNA modifications play a key role in shaping the overall preference of codon usage in proteobacteria. Interestingly, some codons, such as CGA and AGG (encoding arginine), exhibit a surprisingly low level of variation in their frequency of usage, even across genomes with differing GC content. These findings suggest that the evolution of GC content in proteobacterial genomes might be primarily driven by changes in the usage of a specific subset of codons, whose usage is itself influenced by tRNA modifications. [ABSTRACT FROM AUTHOR]

Transfer RNA (tRNA) modifications are essential for the temperature adaptation of thermophilic and psychrophilic organisms as they control the rigidity and flexibility of transcripts. To further understand how specific tRNA modifications are adjusted to maintain functionality in response to temperature fluctuations, we investigated whether tRNA modifications represent an adaptation of bacteria to different growth temperatures (minimal, optimal, and maximal), focusing on closely related psychrophilic (P. halocryophilus and E. sibiricum), mesophilic (B. subtilis), and thermophilic (G. stearothermophilus) Bacillales. Utilizing an RNA sequencing approach combined with chemical pre-treatment of tRNA samples, we systematically profiled dihydrouridine (D), 4-thiouridine (s4U), 7-methyl-guanosine (m7G), and pseudouridine (Ψ) modifications at single-nucleotide resolution. Despite their close relationship, each bacterium exhibited a unique tRNA modification profile. Our findings revealed increased tRNA modifications in the thermophilic bacterium at its optimal growth temperature, particularly showing elevated levels of s4U8 and Ψ55 modifications compared to non-thermophilic bacteria, indicating a temperature-dependent regulation that may contribute to thermotolerance. Furthermore, we observed higher levels of D modifications in psychrophilic and mesophilic bacteria, indicating an adaptive strategy for cold environments by enhancing local flexibility in tRNAs. Our method demonstrated high effectiveness in identifying tRNA modifications compared to an established tool, highlighting its potential for precise tRNA profiling studies. [ABSTRACT FROM AUTHOR]

The Ylistrum japonicum is a commercially valuable scallop known for its long-distance swimming abilities. Despite its economic importance, genetic and genomic research on this species is limited. This study presents the first complete mitochondrial genome of Y. japonicum. The mitochondrial genome is 19,475 bp long and encompasses 13 protein-coding genes, three ribosomal RNA genes, and 23 transfer RNA genes. Two distinct phylogenetic analyses were used to explore the phylogenetic position of the Y. japonicum within the family Pectinidae. Based on one mitochondrial phylogenetic analysis by selecting 15 Pectinidae species and additional outgroup taxa and one single gene phylogenetic analysis by 16S rRNA, two phylogenetic trees were constructed to provide clearer insights into the evolutionary placement of Y. japonicum within the family Pectinidae. Our analysis reveals that Ylistrum is a basal lineage to the Pectininae clade, distinct from its previously assigned tribe, Amusiini. This study offers critical insights into the genetic makeup and evolutionary history of Y. japonicum, enhancing our knowledge of this economically vital species. [ABSTRACT FROM AUTHOR]

Translocation in protein synthesis entails the efficient and accurate movement of the mRNA-[tRNA]2 substrate through the ribosome after peptide bond formation. An essential conformational change during this process is the swiveling of the small subunit head domain about two rRNA 'hinge' elements. Using iterative selection and molecular dynamics simulations, we derive alternate hinge elements capable of translocation in vitro and in vivo and describe their effects on the conformational trajectory of the EF-G-bound, translocating ribosome. In these alternate conformational pathways, we observe a diversity of swivel kinetics, hinge motions, three-dimensional head domain trajectories and tRNA dynamics. By finding alternate conformational pathways of translocation, we identify motions and intermediates that are essential or malleable in this process. These findings highlight the plasticity of protein synthesis and provide a more thorough understanding of the available sequence and conformational landscape of a central biological process. Author summary: Translocation, the motion of the ribosome across its mRNA substrate, is an essential stage of protein synthesis. A key conformational change in this process is the rotation of the ribosome head domain about two rRNA hinges in the direction of translocation, repositioning the mRNA and tRNAs in their final states. Employing iterative selection, we obtain variant hinges capable of performing translocation in vitro and in vivo. Through molecular dynamics simulations, the different variant ribosome translocation conformational trajectories are described. This description reveals different possible conformational pathways to translocation, with varying dynamics, motions and intermediates. The understanding of this conformational malleability can increase our knowledge of protein synthesis function, disruption, evolution, and engineering. [ABSTRACT FROM AUTHOR]

We characterized the complete mitogenome of Nemipterus randalli and performed comparative mitogenomic analysis with previously determined mitochondrial genomes belonging to the family Nemipteridae. Using Sanger sequencing, we identified a 16,642 bp mitogenome containing 37 genes (13 protein-coding, 2 rRNA, 22 tRNA) and a non-coding region. Its gene organization, nucleotide composition, tRNA secondary structure and codon usage are similar to other Nemipteridae mitogenomes. The phylogenetic analysis indicated that Nemipteridae is a monophyletic group. The present study forms the basis for further studies on the population genetics, evolution and phylogeny of N. randalli and its family. [ABSTRACT FROM AUTHOR]

Mosquitoes such as Aedes aegypti must consume a blood meal for the nutrients necessary for egg production. Several transcriptome and proteome changes occur post‐blood meal that likely corresponds with codon usage alterations. Transfer RNA (tRNA) is the adapter molecule that reads messenger RNA codons to add the appropriate amino acid during protein synthesis. Chemical modifications to tRNA enhance codon decoding, improving the accuracy and efficiency of protein synthesis. Here, we examined tRNA modifications and transcripts associated with the blood meal and subsequent periods of vitellogenesis in A. aegypti. More specifically, we assessed tRNA transcript abundance and modification levels in the fat body at critical times post blood‐feeding. Based on a combination of alternative codon usage and identification of particular modifications, we discovered that increased transcription of tyrosine tRNAs is likely critical during the synthesis of egg yolk proteins in the fat body following a blood meal. Altogether, changes in both the abundance and modification of tRNA are essential factors in the process of vitellogenin production after blood‐feeding in mosquitoes. [ABSTRACT FROM AUTHOR]

Genetic code expansion aims to incorporate non‐canonical amino acids (ncAAs) into biological systems, enhancing protein functionality or enabling the in vitro selection of peptides from diverse mRNA displayed libraries. Typically, genetic code expansion has involved reassignment of stop codons to ncAAs through orthogonal translation systems. This review instead focuses on efforts to expand the genetic code by breaking the redundancy of sense codons in vitro and in vivo. In vivo, orthogonal aminoacyl‐tRNA synthetase (AARS)/tRNA/AA systems are able to compete with endogenous machinery, enabling partial to full codon reassignment. Recent approaches, like genome recoding, offer potential solutions to reduce competition. In vitro studies utilize cell extract‐based or reconstituted translation systems, allowing precise control of codon usage via gene design and tRNA addition, making breaking of sense degeneracy easier. In these systems several unsplit codon boxes have been successfully reassigned multiple to ncAAs. These efforts showcase both the successes and challenges in achieving orthogonality and selective codon decoding and point towards a future where the 64 codons can encode more than 30 monomers, enabling new advances in synthetic biology and drug discovery. [ABSTRACT FROM AUTHOR]

T-box riboswitches are noncoding RNA elements involved in genetic regulation of most Gram-positive bacteria. They regulate amino acid metabolism by assessing the aminoacylation status of tRNA, subsequently affecting the transcription or translation of downstream amino acid metabolism-related genes. Here we present single-molecule FRET studies of the Mycobacterium tuberculosis IleS T-box riboswitch, a paradigmatic translational T-box. Results support a two-step binding model, where the tRNA anticodon is recognized first, followed by interactions with the NCCA sequence. Furthermore, after anticodon recognition, tRNA can transiently dock into the discriminator domain even in the absence of the tRNA NCCA-discriminator interactions. Establishment of the NCCA-discriminator interactions significantly stabilizes the fully bound state. Collectively, the data suggest high conformational flexibility in translational T-box riboswitches; and supports a conformational selection model for NCCA recognition. These findings provide a kinetic framework to understand how specific RNA elements underpin the binding affinity and specificity required for gene regulation. smFRET studies of the M. tuberculosis IleS translational T-box riboswitch support a two-step tRNA binding mechanism and the use of conformational selection for tRNA 3' end recognition, which is distinct from the one used by transcriptional T-boxes. [ABSTRACT FROM AUTHOR]

Transfer RNAs (tRNAs) are the most highly modified cellular RNAs, both with respect to the proportion of nucleotides that are modified within the tRNA sequence and with respect to the extraordinary diversity in tRNA modification chemistry. However, the functions of many different tRNA modifications are only beginning to emerge. tRNAs have two general clusters of modifications. The first cluster is within the anticodon stem-loop including several modifications essential for protein translation. The second cluster of modifications is within the tRNA elbow, and roles for these modifications are less clear. In general, tRNA elbow modifications are typically not essential for cell growth, but nonetheless several tRNA elbow modifications have been highly conserved throughout all domains of life. In addition to forming modifications, many tRNA modifying enzymes have been demonstrated or hypothesized to also play an important role in folding tRNA acting as tRNA chaperones. In this review, we summarize the known functions of tRNA modifying enzymes throughout the lifecycle of a tRNA molecule, from transcription to degradation. Thereby, we describe how tRNA modification and folding by tRNA modifying enzymes enhance tRNA maturation, tRNA aminoacylation, and tRNA function during protein synthesis, ultimately impacting cellular phenotypes and disease. [ABSTRACT FROM AUTHOR]

Archaeosine (G+) is an archaea-specific tRNA modification synthesized via multiple steps. In the first step, archaeosine tRNA guanine transglucosylase (ArcTGT) exchanges the G15 base in tRNA with 7-cyano-7-deazaguanine (preQ0). In Euryarchaea, preQ015 in tRNA is further modified by archaeosine synthase (ArcS). Thermococcus kodakarensis ArcS catalyzes a lysine-transfer reaction to produce preQ0-lysine (preQ0-Lys) as an intermediate. The resulting preQ0-Lys15 in tRNA is converted to G+15 by a radical S-adenosyl-L-methionine enzyme for archaeosine formation (RaSEA), which forms a complex with ArcS. Here, we focus on the substrate tRNA recognition mechanism of ArcS. Kinetic parameters of ArcS for lysine and tRNA-preQ0 were determined using a purified enzyme. RNA fragments containing preQ0 were prepared from Saccharomyces cerevisiae tRNAphe-preQ015. ArcS transferred 14C-labeled lysine to RNA fragments. Furthermore, ArcS transferred lysine to preQ0 nucleoside and preQ0 nucleoside 50 -monophosphate. Thus, the L-shaped structure and the sequence of tRNA are not essential for the lysine-transfer reaction by ArcS. However, the presence of D-arm structure accelerates the lysine-transfer reaction. Because ArcTGT from thermophilic archaea recognizes the common D-arm structure, we expected the combination of T. kodakarensis ArcTGT and ArcS and RaSEA complex would result in the formation of preQ0-Lys15 in all tRNAs. This hypothesis was confirmed using 46 T. kodakarensis tRNA transcripts and three Haloferax volcanii tRNA transcripts. In addition, ArcTGT did not exchange the preQ0-Lys15 in tRNA with guanine or preQ0 base, showing that formation of tRNA-preQ0-Lys by ArcS plays a role in preventing the reverse reaction in G+biosynthesis. [ABSTRACT FROM AUTHOR]

The targeted capture of ultraconserved elements (UCEs) has substantially increased the amount of genetic data available for phylogenomic reconstructions. These capture datasets frequently contain mitochondrial DNA as a by‐product, often in the form of complete mitogenomes. These can be efficiently harvested to expand existing datasets without additional costs. Here, we present new mitochondrial genomes for six marine angelfish species (F: Pomacanthidae), assembled and annotated from off‐target UCE reads. We provide the first comparative analysis of all mitochondrial genomes available for the Pomacanthidae. Results showed that the average length of pomacanthid mitogenomes is 16.8 kbp. Total GC and AT content varied between 44.5% and 46.3%, and 53.7% and 55.5%, respectively. The architecture of angelfish mitogenomes was comparable to that seen in other fish species with 13 protein‐coding genes (PCGs), 22 transfer RNA genes, two ribosomal RNA genes and the control region. All 13 PCGs evolved under purifying selection, highlighting a high level of selection pressure and gene expression to preserve genetic integrity. The ND6 and ATP8 genes had the highest ratio of non‐synonymous (dN) to synonymous (dS) substitutions, indicating a relaxation of purifying selection constraints. Finally, these newly assembled mitogenomes will allow further investigations of the population genetics, systematics and evolutionary biology of one of the most prominent reef fish family in the aquarium trade. [ABSTRACT FROM AUTHOR]

Simple Summary: Fetal macrosomia is when a baby's weight at birth is equal to or greater than 4000 g or 4500 g. The rising incidence of macrosomia poses a significant challenge in obstetrics, as it can have serious health consequences for both mothers and babies. The maternal gut microbiome can influence the health of pregnant women and their babies, with potential effects on birth weight. However, research on the link between the microbiome and birth weight, especially macrosomia, is limited; further investigation is needed. Here, we discovered a connection between the maternal gut microbiome and macrosomia. Our findings present novel opportunities for preventing macrosomia by manipulating the composition of the intestinal microbiota. Early prediction models using gut microbiota and clinical indicators can predict macrosomia. Fetal macrosomia is defined as a birthweight ≥4000 g and causes harm to pregnant women and fetuses. Studies reported that the maternal intestinal microbiome plays a key role in the establishment, growth, and development of the fetal intestinal microbiome. However, whether there is a relationship between maternal gut microbiota and macrosomia remains unclear. Our study aimed to identify gut microbiota that may be related to the occurrence of macrosomia, explore the possible mechanisms by which it causes macrosomia, and establish a prediction model to determine the feasibility of predicting macrosomia by early maternal gut microbiota. We conducted a nested case-control study based on an early pregnancy cohort (ChiCTR1900020652) in the Maternity and Child Health Hospital of Hunan Province on fecal samples of 93 women (31 delivered macrosomia as the case group and 62 delivered normal birth weight newborns as the control group) collected and included in this study. We performed metagenomic analysis to compare the composition and function of the gut microbiome between cases and controls. Correlation analysis was used to explore the association of differential species and differential functional pathways. A random forest model was used to construct an early pregnancy prediction model for macrosomia. At the species level, there were more Bacteroides salyersiae, Bacteroides plebeius, Ruminococcus lactaris, and Bacteroides ovatus in the intestinal microbiome of macrosomias' mothers compared with mothers bearing fetuses that had normal birth weight. Functional pathways of the gut microbiome including gondoate biosynthesis, L-histidine degradation III, cis-vaccenate biosynthesis, L-arginine biosynthesis III, tRNA processing, and mannitol cycle, which were more abundant in the macrosomia group. Significant correlations were found between species and functional pathways. Bacteroides plebeius was significantly associated with the pathway of cis-vaccenate biosynthesis (r = 0.28, p = 0.005) and gondoate biosynthesis (r = 0.28, p < 0.001) and Bacteroides ovatus was positively associated with the pathway of cis-vaccenate biosynthesis (r = 0.29, p = 0.005) and gondoate biosynthesis (r = 0.32, p = 0.002). Bacteroides salyersiae was significantly associated with the pathway of cis-vaccenate biosynthesis (r = 0.24, p = 0.018), gondoate biosynthesis (r = 0.31, p = 0.003), and L–histidine degradation III (r = 0.22, p = 0.291). Finally, four differential species and four clinical indicators were included in the random forest model for predicting macrosomia. The areas under the working characteristic curves of the training and validation sets were 0.935 (95% CI: 0.851~0.979) and 0.909 (95% CI: 0.679~0.992), respectively. Maternal gut microbiota in early pregnancy may play an important role in the development of macrosomia and can be used as potential predictors to prevent macrosomia. [ABSTRACT FROM AUTHOR]

Key message: We reported the graph-based mitochondrial genomes of three foundation species (Saccharum spontaneum, S. robustum and S. officinarum) for the first time. The results revealed pan-structural variation and evolutionary processes in the mitochondrial genomes within Saccharum. Saccharum belongs to the Andropogoneae, and cultivars species in Saccharum contribute nearly 80% of sugar production in the world. To explore the genomic studies in Saccharum, we assembled 15 complete mitochondrial genomes (mitogenome) of three foundation species (Saccharum spontaneum, S. robustum and S. officinarum) using Illumina and Oxford Nanopore Technologies sequencing data. The mitogenomes of the three species were divided into a total of eight types based on contig numbers and linkages. All mitogenomes in the three species encoded 51 unique genes, including 32 protein-coding, 3 ribosomal RNA (rRNA) and 16 transfer RNA (tRNA) genes. The existence of long and short-repeat-mediated recombinations in the mitogenome of S. officinarum and S. robustum was revealed and confirmed through PCR validation. Furthermore, employing comparative genomics and phylogenetic analyses of the organelle genomes, we unveiled the evolutionary relationships and history of the major interspecific lineages in Saccharum genus. Phylogenetic analyses of homologous fragments between S. officinarum and S. robustum showed that S. officinarum and S. robustum are phylogenetically distinct and that they were likely parallel rather than domesticated. The variations between ancient (S. sinense and S. barberi) and modern cultivated species (S. hybrid) possibly resulted from hybridization involving different S. officinarum accessions. Lastly, this project reported the first graph-based mitogenomes of three Saccharum species, and a systematic comparison of the structural organization, evolutionary processes, and pan-structural variation of the Saccharum mitogenomes revealed the differential features of the Saccharum mitogenomes. [ABSTRACT FROM AUTHOR]

The article explores the essential functions of iron-sulfur (Fe-S) clusters in a wide range of biological processes, including their assembly and biogenesis, and their crucial role in cellular functions. It highlights the involvement of Fe-S clusters in human health, particularly their links to diseases such as ALS, cancers, and diabetes, while also examining their significance in cellular homeostasis, oxygen sensing, and viral pathogenesis.

Background: Gestational Diabetes Mellitus (GDM) poses significant risks to maternal and fetal health. Current diagnostic methods based on glucose tolerance tests have limitations for early detection. tRNA‐derived small RNAs (tsRNAs) have emerged as potential molecular regulators in various diseases, including metabolic disorders. However, the diagnostic value of tsRNAs in plasma for early GDM or postpartum remains unclear. Methods: This longitudinal study profiled the expression of tsRNAs across different gestational stages and postpartum in women with GDM (n = 40) and healthy control gestational women (HCs, n = 40). High‐throughput small RNA sequencing identified candidate tsRNAs, which were then validated and correlated with clinical biochemical markers such as fasting blood glucose (FBG), HOMA‐IR, and GHbA1c. Results: tRF‐1:32‐Val‐AAC‐1‐M6, tRF‐1:31‐Glu‐CTC‐1‐M2, and tRF‐1:30‐Gly‐CCC‐1‐M4 were consistently upregulated in the GDM group compared to HCs during the second trimester (p < 0.05). Only tRF‐1:31‐Glu‐CTC‐1‐M2 was highly expressed during the first trimester, and tRF‐1:30‐Gly‐CCC‐1‐M4 increased during postpartum. tRF‐1:31‐Glu‐CTC‐1‐M2 showed a significant correlation with FBG levels in the first trimester (R = 0.317, p = 0.047). The expression of tRF‐1:30‐Gly‐CCC‐1‐M4 was significantly correlated with HOMA‐IR (r = 0.65, p < 0.001) and GHBA1c (r = 0.33, p = 0.037) during postpartum. A joint diagnostic model incorporating tsRNAs expression and clinical markers demonstrated enhanced predictive power for GDM (ROC AUC = 0.768). Conclusion: Our results revealed distinct expression patterns of specific tsRNAs in GDM, showcasing their correlation with key metabolic parameters. This underscores their promising role as biomarkers for early prediction and diagnosis of GDM. The integration of tRFs into a composite biomarker panel holds the potential to improve clinical outcomes by enabling personalized risk assessment and targeted interventions. [ABSTRACT FROM AUTHOR]

The amide proteogenic amino acids, asparagine and glutamine, are two of the twenty amino acids used in translation by all known life. The aminoacyl‐tRNA synthetases for asparagine and glutamine, asparaginyl‐tRNA synthetase and glutaminyl tRNA synthetase, evolved after the split in the last universal common ancestor of modern organisms. Before that split, life used two‐step indirect pathways to synthesize asparagine and glutamine on their cognate tRNAs to form the aminoacyl‐tRNA used in translation. These two‐step pathways were retained throughout much of the bacterial and archaeal domains of life and eukaryotic organelles. The indirect routes use non‐discriminating aminoacyl‐tRNA synthetases (non‐discriminating aspartyl‐tRNA synthetase and non‐discriminating glutamyl‐tRNA synthetase) to misaminoacylate the tRNA. The misaminoacylated tRNA formed is then transamidated into the amide aminoacyl‐tRNA used in protein synthesis by tRNA‐dependent amidotransferases (GatCAB and GatDE). The enzymes and tRNAs involved assemble into complexes known as transamidosomes to help maintain translational fidelity. These pathways have evolved to meet the varied cellular needs across a diverse set of organisms, leading to significant variation. In certain bacteria, the indirect pathways may provide a means to adapt to cellular stress by reducing the fidelity of protein synthesis. The retention of these indirect pathways versus acquisition of asparaginyl‐tRNA synthetase and glutaminyl tRNA synthetase in lineages likely involves a complex interplay of the competing uses of glutamine and asparagine beyond translation, energetic costs, co‐evolution between enzymes and tRNA, and involvement in stress response that await further investigation. [ABSTRACT FROM AUTHOR]

Many aspects of the life cycle and genetics of (+)RNA viruses are directed by their genome replication organelles (ROs), which form as spherular membrane invaginations (this review's primary focus) or double-membrane vesicles. Recent cryo-electron microscopy (cryo-EM) studies have transformed understanding of RO structure and assembly. RO membrane vesicles containing dsRNA replication intermediates are gated to the cytosol by ringed 'crown' complexes of viral RNA replication proteins that direct synthesis, 5′ capping, and release of new mRNA-sense genomic and subgenomic RNAs. Although nodaviruses and alphaviruses are only distantly related, their RO crowns conserve 12-fold symmetry, key enzymatic and structural motifs, and multistage assembly. Continuing studies should enable new antiviral controls and reveal how crown assembly steps are coupled to viral (−)RNA and (+)RNA synthesis, RO vesicle formation, and progeny RNA transfer to translation, new RO formation, and encapsidation. Positive-strand RNA [(+)RNA] viruses include pandemic SARS-CoV-2, tumor-inducing hepatitis C virus, debilitating chikungunya virus (CHIKV), lethal encephalitis viruses, and many other major pathogens. (+)RNA viruses replicate their RNA genomes in virus-induced replication organelles (ROs) that also evolve new viral species and variants by recombination and mutation and are crucial virus control targets. Recent cryo-electron microscopy (cryo-EM) reveals that viral RNA replication proteins form striking ringed 'crowns' at RO vesicle junctions with the cytosol. These crowns direct RO vesicle formation, viral (−)RNA and (+)RNA synthesis and capping, innate immune escape, and transfer of progeny (+)RNA genomes into translation and encapsidation. Ongoing studies are illuminating crown assembly, sequential functions, host factor interactions, etc., with significant implications for control and beneficial uses of viruses. [ABSTRACT FROM AUTHOR]

In the yeast genera Saccharomycopsis and Ascoidea , which comprise the taxonomic order Ascoideales, nuclear genes use a nonstandard genetic code in which CUG codons are translated as serine instead of leucine, due to a tRNA-Ser with the unusual anticodon CAG. However, some species in this clade also retain an ancestral tRNA-Leu gene with the same anticodon. One of these species, Ascoidea asiatica , has been shown to have a stochastic proteome in which proteins contain ∼50% Ser and 50% Leu at CUG codon sites, whereas previously examined Saccharomycopsis species translate CUG only as Ser. Here, we investigated the presence, conservation, and possible functionality of the tRNA-Leu(CAG) gene in the genus Saccharomycopsis. We sequenced the genomes of 23 strains that, together with previously available data, include almost every known species of this genus. We found that most Saccharomycopsis species have genes for both tRNA-Leu(CAG) and tRNA-Ser(CAG). However, tRNA-Leu(CAG) has been lost in Saccharomycopsis synnaedendra and Saccharomycopsis microspora , and its predicted cloverleaf structure is aberrant in all the other Saccharomycopsis species. We deleted the tRNA-Leu(CAG) gene of Saccharomycopsis capsularis and found that it is not essential. Proteomic analyses in vegetative and sporulating cultures of S. capsularis and Saccharomycopsis fermentans showed only translation of CUG as Ser. Despite its unusual structure, the tRNA-Leu(CAG) gene shows evidence of sequence conservation among Saccharomycopsis species, particularly in its acceptor stem and leucine identity elements, which suggests that it may have been retained in order to carry out an unknown nontranslational function. [ABSTRACT FROM AUTHOR]

Gastrochilus is an orchid genus containing about 70 species in tropical and subtropical Asia with high morphological diversity. The phylogenetic relationships among this genus have not been fully resolved, and the plastome evolution has not been investigated either. In this study, five plastomes of Gastrochilus were newly reported, and sixteen plastomes of Gastrochilus were used to conduct comparative and phylogenetic analyses. Our results showed that the Gastrochilus plastomes ranged from 146,183 to 148,666 bp, with a GC content of 36.7–36.9%. There were 120 genes annotated, consisting of 74 protein-coding genes, 38 tRNA genes, and 8 rRNA genes. No contraction and expansion of IR borders, gene rearrangements, or inversions were detected. Additionally, the repeat sequences and codon usage bias of Gastrochilus plastomes were highly conserved. Twenty hypervariable regions were selected as potential DNA barcodes. The phylogenetic relationships within Gastrochilus were well resolved based on the whole plastome, especially among main clades. Furthermore, both molecular and morphological data strongly supported Haraella retrocalla as a member of Gastrochilus (G. retrocallus). [ABSTRACT FROM AUTHOR]

Background: Breast cancer (BC) is the most prevalent cancer among women worldwide. Although advances have been made in the identification of predictive biomarkers, current options for early diagnosis and prognostic analysis are still suboptimal. Recently, transfer-RNA-derived RNA fragments (tRFs) have emerged as a class of small noncoding RNAs that play a role in the cancer progression. The authors aimed to identify a specific class of tRFs as a molecular marker for BC diagnosis and prognosis in clinical management. Methods: The levels of 5′-tRF-His-GTG were quantified in BC tissue (n = 101) and inflammatory normal breast tissue (n = 22) using in situ hybridization. Clinicopathological parameters were obtained, including age, tumor node metastasis stage, hormone receptor status, histopathological grade, lymphovascular invasion, and recurrence. The correlation between the expression of 5′-tRF-His-GTG and these parameters in different BC subtypes was analyzed. Patient death and cancer progression were regarded as clinical endpoints in the survival analysis. Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were also performed to predict the involvement in pivotal biological process. Results: The expression of 5′-tRF-His-GTG was significantly downregulated in BC tissues and was in connection with T stage in human epidermal growth factor 2-positive and basal-like BC, as well as N stage and histopathological grade in luminal BC. Patients with low expression of 5′-tRF-His-GTG had a poor overall survival rate. Statistics of GO and KEGG pathway revealed that cation channel activity, protein catabolic process, response to temperature stimulus, cell cycle, focal adhesion, and glycerophospholipid metabolism were significantly enriched. Conclusions: This study suggests that the assessment of 5′-tRF-His-GTG expression could serve as a novel biomarker for individual diagnosis and prognosis in BC. [ABSTRACT FROM AUTHOR]

The inconsistency between traditional morphological taxonomy and molecular phylogenetic data is a major issue that puzzles the study of fish classification and evolution. Although mitochondrial genes are commonly used in phylogenetic analyses to compare fish species, the mitochondrial evolution and diversity of Schistura are still not well understood. To better understand the evolution of Schistura, we sequenced the mitochondrial genome of Schistura fasciolata and compared it with other species of Schistura. A 16,588 bp circular mitochondrial genome of S. fasciolata was obtained and it contains 13 protein-coding, 22 transfer RNA, and two ribosomal RNA genes, and a non-coding control region. The gene arrangement in the mitochondrial genomes of all Schistura species was consistent. However, we also found that S. fasciolata was not monophyletic. Although mitochondrial genes can be effectively used for Schistura species identification, they may not be suitable for inferring the evolutionary process of Schistura species. These results provide support for the use of mitochondrial genes in identifying Schistura species, and also serve as a warning against mistakenly using them to evaluate the evolution process of Schistura species. [ABSTRACT FROM AUTHOR]

The capybara, Hydrochoerus hydrochaeris, is a rodent that inhabits the wetlands of South America. Despite being widely distributed and having the nuclear genomes available, there has been a lack of information regarding the mitochondrial genome. To address this, we conducted the assembly and annotation of the capybara's mitochondrial genome and reconstructed the phylogeny of Parvorder Caviomorpha. The assembly was conducted under the de novo method with GetOrganelle, while the annotation was performed with MitoZ. Analysis of relative codon synonymous usage was conducted in the mitochondrial genomes of the capybara and two other Caviidae species: Cavia porcellus and Cavia aperea. The phylogenetic inference was conducted under the Maximum Likelihood method, using the 13 protein-coding genes, including the capybara as well as other 42 Caviomorpha mitochondrial genomes. The resulting capybara mitochondrial genome consisted of 16,681 bp, 37 genes (22 tRNAs, 2 rRNAs, and 13 protein-coding genes) and a control region. The number of genes and their arrangement corresponds to the pattern observed in most mammalian species. The capybara was recovered as a sister group of Cavia. Caviidae was reconstructed as a monophyletic group that is closely related to Cuniculidae. Overall, this study provides valuable insights into the mitochondrial evolution and evolutionary relationships of the capybara. [ABSTRACT FROM AUTHOR]

Methyltransferase-like 23 (METTL23) is a kind of RNA methyltransferase that catalyzes the methylation transfer to the N6-adenosine of RNA, serving as one of the key mediators in this process. However, the METTL23 gene has been poorly researched in pigs. In this study, we investigated the genetic effects of METTL23 single-nucleotide polymorphism(SNPs) on reproductive traits in Kele pigs. The DNA was extracted from 228 healthy multiparous Kele sows, and Sanger sequencing revealed three SNPs, g.4804958 G > T (intron 2), g.4805082 C > T (exon 2), and g.4806821 A > G (exon 3). The polymorphism information content (PIC) for each SNP was 0.264, 0.25, and 0.354, indicating moderate polymorphism (0.25 < PIC < 0.5) and providing genetic information. Linkage disequilibrium analysis showed no strong linkage disequilibrium between the three SNPs. The association analysis revealed that in the SNP g.4804958 G > T individuals with the GG genotype had a significantly higher number of piglets born alive, litter birth weight, number of weaned piglets, and weaning litter weight compared to those with the TT genotype (p < 0.05). Individuals with the GG genotype in the SNP g.4806821 A > G group had significantly higher litter birth weight and average birth weight than those with the AA genotype (p < 0.05). The H4H4 diplotype showed significant effects on the number of piglets born alive, litter birth weight, number of weaned piglets, weaning litter weight, and weaning weight (p < 0.05). Together, the METTL23 gene could be used as a candidate gene for the selection of reproductive traits in Kele pigs. [ABSTRACT FROM AUTHOR]

Hepatocellular adenoma (HCA) and hepatocellular carcinoma (HCC) can be difficult to differentiate but must be diagnosed correctly as treatment and prognosis for these tumors differ markedly. Relevant diagnostic biomarkers are thus needed, and those identified in dogs may have utility in human medicine because of the similarities between human and canine HCA and HCC. A tRNA-derived fragment (tRF), tRNA-Val, is a promising potential biomarker for canine mammary gland tumors but has not previously been investigated in hepatic tumors. Accordingly, we aimed to elucidate the potential utility of tRNA-Val as a biomarker for canine HCA and HCC using clinical samples (tumor tissue and plasma extracellular vesicles [EVs]) and tumor cell lines with qRT-PCR assays. We also investigated relevant functions and signaling pathways with bioinformatic analyses (Gene Ontology and Kyoto Encyclopedia of Genes and Genomes). tRNA-Val was markedly downregulated in HCC tumor tissue versus HCA tumor tissue and normal liver tissue, and a similar trend was shown in plasma EVs and HCC cell lines versus healthy controls. Based on areas under the receiver operating characteristic curves (AUCs), tRNA-Val significantly distinguished HCC (AUC = 1.00, p = 0.001) from healthy controls in plasma EVs and HCC from HCA (AUC = 0.950, p = 0.01). Bioinformatics analysis revealed that tRNA-Val may be primarily involved in DNA repair, mRNA processing, and splicing and may be linked to the N-glycan and ubiquitin-mediated proteasome pathways. This is the first report on the expression of tRNA-Val in canine HCC and HCA and its possible functions and signaling pathways. We suggest that tRNA-Val could be a promising novel biomarker to distinguish canine HCC from HCA. This study provides evidence for a greater understanding of the role played by tRNA-Val in the development of canine HCC. [ABSTRACT FROM AUTHOR]

A Ti3C2TxMXene-based biosensor has been developed and the photocatalytic atom transfer radical polymerization (photo ATRP) amplification strategy applied to detect target miRNA-21 (tRNA). Initially, Ti3C2TxMXene nanosheets were synthesized from the Ti3AlC2 MAX precursor via selective aluminum etching. Then, functionalization of Ti3C2TxMXene nanosheets with 3-aminopropyl triethoxysilane (APTES) via silylation reactions to facilitate covalent bonding with hairpin DNA biomolecules specifically designed for tRNA detection. Upon binding with the tRNA, the hairpin DNA liberated the azide (N₃) group, initiating a click reaction to affix to the photo ATRP initiator. Through the ATRP photoreaction, facilitated by an organic photoredox catalyst and light, a significant amount of ferrocenyl methyl methacrylate (FMMA) monomer was immobilized on the electrode. Therefore, the electrochemical signal is amplified. The electrochemical efficacy of the biosensor was assessed using square wave voltammetry (SWV). Under optimized conditions, the biosensor demonstrated remarkable sensitivity in detecting tRNA, with a linear detection range from 0.01 fM to 10 pM and a detection limit of 2.81 aM. The findings elucidate that the developed biosensor, in conjunction with the photo ATRP strategy, offers reproducibility, stability, and increased sensitivity, underscoring its potential applications within the experimental medical sector of the biomolecular industry. [ABSTRACT FROM AUTHOR]

Triticummilitinae Zhuk. et Migusch., a tetraploid wheat with the GAA genome, is considered a natural naked mutant of Triticum timopheevii Zhuk. Previously, the karyotype and crossing characteristics of this wheat were examined. To clarify the origin and relationships of T. militinae with other representatives of the wheat family, analysis of its chloroplast genome, which remained unexplored, is of great interest. In the present study, for the first time, sequencing and annotation of the complete chloroplast genome of T. militinae, the size of which was found to be 135 898 bp, was conducted. The plastome of this wheat is composed of two inverted repeats, each of 21 552 bp in length, the small single-copy (SSC) region of 12 791 bp, and the large single-copy (LSC) region of 80 003 bp. The chloroplast genome of T. militinae contains 132 annotated structural genes, of which 85 genes are protein-coding, 31 are tRNA genes, and four genes code for rRNA. [ABSTRACT FROM AUTHOR]

Thirteen-lined ground squirrels (TLGSs) are obligate hibernators that cycle between torpor (low metabolic rate and body temperature) and interbout euthermia (IBE; typical euthermic body temperature and metabolism) from late autumn to spring. Many physiological changes occur throughout hibernation, including a reduction in liver mitochondrial metabolism during torpor, which is reversed during arousal to interbout euthermia. Nuclear-encoded microRNA (miRNA, small posttranscriptional regulator molecules) differ in abundance throughout TLGS hibernation and have been shown to regulate mitochondrial gene expression in mammalian cell culture (where they are referred to as mitomiRs). This study characterized differences in mitomiR profiles from TLGS liver mitochondria isolated during summer, torpor, and IBE, and predicted their mitochondrial targets. Using small RNA sequencing, differentially abundant mitomiRs were identified between hibernation states, and using quantitative PCR analysis, we quantified the expression of predicted mitochondrial mRNA targets. Most differences in mitomiR abundances were seasonal (i.e., between summer and winter) with only one mitomiR differentially abundant between IBE and torpor. Multiple factor analysis (MFA) revealed three clusters divided by hibernation states, where clustering was predominantly driven by mitomiR abundances. Nine of these differentially abundant mitomiRs had predicted mitochondrial RNA targets, including subunits of electron transfer system complexes I and IV, 12S rRNA, and two tRNAs. Overall, mitomiRs were predicted to suppress the expression of their mitochondrial targets and may have some involvement in regulating protein translation in mitochondria. This study found differences in mitomiR abundances between seasons and hibernation states of TLGS and suggests potential mechanisms for regulating the mitochondrial electron transfer system. NEW & NOTEWORTHY: During the hibernation season, thirteen-lined ground squirrels periodically increase metabolism remarkably between torpor and interbout euthermia (IBE). This process involves rapid reactivation of mitochondrial respiration. We predicted that mitochondrial microRNA (mitomiRs) might be altered during this response. We found that the abundance of 38 liver mitomiRs differs based on hibernation state (summer, IBE, and torpor). Small RNA sequencing identified mitomiR profiles, including some mitomiRs that are predicted to bind to mitochondrial RNAs. [ABSTRACT FROM AUTHOR]

Environmental stressors can induce paternal epigenetic modifications that are a key determinant of the intergenerational inheritance of acquired phenotypes in mammals. Some of them can affect phenotypic expression through inducing changes in tRNA-derived small RNAs (tsRNAs), which modify paternal epigenetic regulation in sperm. However, it is unclear how these stressors can affect changes in the expression levels of tsRNAs and their related endonucleases in the male reproductive organs. We found that Ribonuclease inhibitor 1 (RNH1), an oxidation responder, interacts with ANG to regulate sperm tsRNA generation in the mouse caput epididymis. On the other hand, inflammation and oxidative stress induced by either lipopolysaccharide (LPS) or palmitate (PA) treatments weakened the RNH1-ANG interaction in the epididymal epithelial cells (EEC). Accordingly, ANG translocation increased from the nucleus to the cytoplasm, which led to ANG upregulation and increases in cytoplasmic tsRNA expression levels. In conclusion, as an antioxidant, RNH1 regulates tsRNA generation through targeting ANG in the mouse caput epididymis. Moreover, the tsRNA is an epigenetic factor in sperm that modulates paternal inheritance in offspring via the fertilization process. [ABSTRACT FROM AUTHOR]

The rules of the genetic code are implemented by the unique features that define the amino acid identity of each transfer RNA (tRNA). These features, known as "identity elements," mark tRNAs for recognition by aminoacyl-tRNA synthetases (ARSs), the enzymes responsible for ligating amino acids to tRNAs. While tRNA identity elements enable stringent substrate selectivity of ARSs, these enzymes are prone to errors during amino acid selection, leading to the synthesis of incorrect aminoacyl-tRNAs that jeopardize the fidelity of protein synthesis. Many error-prone ARSs have evolved specialized domains that hydrolyze incorrectly synthesized aminoacyl-tRNAs. These domains, known as editing domains, also exist as free-standing enzymes and, together with ARSs, safeguard protein synthesis fidelity. Here, we discuss how the same identity elements that define tRNA aminoacylation play an integral role in aminoacyl-tRNA editing, synergistically ensuring the correct translation of genetic information into proteins. Moreover, we review the distinct strategies of tRNA selection used by editing enzymes and ARSs to avoid undesired hydrolysis of correctly aminoacylated tRNAs. [ABSTRACT FROM AUTHOR]

Colias sifanica Grum-Grshimailo, 1891, is a typical montane butterfly species which occurs on the Qinghai-Tibet Plateau (QTP) and adjacent regions in China. In this study, the complete mitochondrial genome of this species was assembled from data generated by next-generation sequencing. The mitogenome was 15,151 bp in length and comprised 13 protein-coding genes (PCGs), 2 ribosomal RNA genes, 22 transfer RNA genes and a control region. The base composition of the C. sifanica mitogenome was 39.7% A, 41.3% T, 11.3% C and 7.7% G, significantly AT biased as commonly found in other Pieridae mitogenomes. Phylogenetic analyses based on all PCGs using both the maximum likelihood and Bayesian inference methods indicated that C. sifanica is closely related to C. fieldii with high support values, and the phylogenetic relationship of (Dercas + ((Gandaca + Gonepteryx) + (Phoebis + (Anteos + (Catopsilia + (Zerene + Colias))))))) was shown for the tribe Coliadini. Though both the mitogenomic gene order and overall base composition were found to be conserved, different Ka/Ks ratios for several mitogenomic PCGs were detected between Colias and other species in the tribe Coliadini. [ABSTRACT FROM AUTHOR]

Pyankovia brachiata (Pall.) Akhani & Roalson 2007, is an annual plant belonging to the genus Pyankovia, family Amaranthaceae, which is widely distributed in the inland deserts of Northwest China. P. brachiata was previously categorized under the genus Salsola in Salsoleae and has been a long-standing topic of debate. Therefore, the complete chloroplast genome of P. brachiata must be studied to provide a theoretical reference for species classification. In this study, we sequenced P. brachiata samples and determined the species' complete chloroplast genome. The complete chloroplast genome was 149,922 bp in length, with one large single copy (LSC: 83,565 bp), one small single copy (SSC: 18,535 bp), and two inverted repeat regions (IRa and IRb, 23,911 bp each). It contains 132 genes, including 87 protein-coding, eight rRNA, and 37 tRNA genes. The phylogenetic position showed that P. brachiata has the closest relationship with Caroxylon passerinum (accession number: NC057191.1). This study will provide genetic information and be beneficial to understanding the systematic position of P. brachiata within the Amaranthaceae. [ABSTRACT FROM AUTHOR]

Camellia flava (Pit.) Sealy 1949 is a rare and precious species with golden flowers, which hold important ornamental and breeding values. In this study, the complete chloroplast genome of C. flava is reported for the first time. The chloroplast genome exhibits a typical quadripartite structure with a total length of 156,670 bp and a GC content of 37.32%, including a large single-copy region (86,250 bp), a small single-copy region (18,292 bp), and a pair of inverted repeat regions (26,064 bp). A total of 133 genes, including 88 protein-coding genes, 37 tRNA genes, and 8 rRNA genes were annotated. The phylogenetic analysis revealed a close relationship between C. flava and C. tamdaoensis. The chloroplast genome sequence of C. flava serves as a valuable resource for further breeding research and genetic phylogenetic studies. [ABSTRACT FROM AUTHOR]

Lindernia crustacea (L.) F. Muell 1882, a species in the Linderniaceae family, holds traditional medicinal value in China. To investigate its genetic diversity, we assembled, annotated, and characterized the first complete chloroplast genome of L. crustacea using Illumina sequencing data and various bioinformatics tools. The genome is 153,647 bp in length, with a GC content of 37.6%. It exhibits a typical quadripartite structure, consisting of a large single-copy region (LSC) of 85,411 bp, a small single-copy region (SSC) of 18,724 bp, and two inverted repeat sequences (IRa and IRb) of 25,816 bp each. The genome was predicted to contain 131 genes, including 87 protein-coding genes, 36 tRNA genes, and eight rRNA genes. Phylogenomic analysis indicated that L. crustacea is closely related to L. stricta. These findings provide a foundation for further research on the evolution and potential medicinal applications of the Linderniaceae family. [ABSTRACT FROM AUTHOR]