Bombyx mori Tryptophanyl-tRNA synthetase (BmWARS) belongs to the family of Ic-like aminoacyl-tRNA synthetases (aaRSs), whose specific recognition of the substrate Trp, tRNA, maintains the fidelity of protein synthesis. In this study, BmWARS was cloned and characterized from the midgut of the silkworm, Bombyx mori, resulting in an open reading frame (ORF) with a full length of 1,149 bp, which can encode 382 Aa. BmWARS is localized in the cytoplasm, and is expressed in all tissues of the silkworm, with higher expression in the testis, ovary, silk gland and malpighian tubule. The expression of BmWARS was significantly up-regulated in the midgut and silk gland after infection with Bombyx mori nuclear polyhedrosis virus (BmNPV). In addition, overexpression of BmWARS inhibited BmNPV infection and replication extremely significantly, while interference with BmWARS expression promoted BmNPV infection and replication. Analysis of the immune pathways in which BmWARS may be involved revealed that the expression of the key genes of the PI3K-Akt pathway, BmPI3K, BmAkt, BmPDK1, BmeIF4E, BmS6, and p-Akt protein was significantly reduced, whereas the expression of BmPTEN, BmFoxO, and BmCaspase9 was significantly increased in the cells that overexpressed BmWARS and were infected with BmNPV. Meanwhile, the results of the study interfering with the expression of BmWARS were completely opposite to those of the study overexpressing BmWARS. This is the first report that BmWARS has antiviral effects in Bombyx mori. Moreover, BmWARS inhibits BmNPV infection and replication in Bombyx mori cells by promoting apoptosis and inhibiting cell proliferation.

Interstitial lung disease (ILD) consists of a group of immune-mediated disorders that can cause inflammation and progressive fibrosis of the lungs, representing an area of unmet medical need given the lack of disease-modifying therapies and toxicities associated with current treatment options. Tissue-specific splice variants (SVs) of human aminoacyl-tRNA synthetases (aaRSs) are catalytic nulls thought to confer regulatory functions. One example from human histidyl-tRNA synthetase (HARS), termed HARSWHEP because the splicing event resulted in a protein encompassing the WHEP-TRS domain of HARS (a structurally conserved domain found in multiple aaRSs), is enriched in human lung and up-regulated by inflammatory cytokines in lung and immune cells. Structural analysis of HARSWHEP confirmed a well-organized helix-turn-helix motif. This motif bound specifically and selectively to neuropilin-2 (NRP2), a receptor expressed by myeloid cells in active sites of inflammation, to inhibit expression of proinflammatory receptors and cytokines and to down-regulate inflammatory pathways in primary human macrophages. In animal models of lung injury and ILD, including bleomycin treatment, silicosis, sarcoidosis, chronic hypersensitivity pneumonitis, systemic sclerosis, and rheumatoid arthritis-ILD, HARSWHEP reduced lung inflammation, immune cell infiltration, and fibrosis. In patients with sarcoidosis, efzofitimod treatment resulted in down-regulation of gene expression for inflammatory pathways in peripheral immune cells and stabilization of inflammatory biomarkers in serum after steroid tapering. We demonstrate the immunomodulatory activity of HARSWHEP and present preclinical data supporting ongoing clinical development of the biologic efzofitimod based on HARSWHEP in ILD.

Antimicrobial peptides (AMPs), derived from a variety of proteins such as ribosomal proteins, play a pivotal role in the innate immune system. However, information regarding ribosomal protein-derived AMPs is currently limited and their mechanisms of action remain poorly defined. Here we identified and characterized the antibacterial activity of amphioxus RPL27 (BjRPL27) and its core functional region located at residues 51-72 (termed BjRPL2751-72). We found that BjRPL27 expression was upregulated in the hepatic caecum following bacterial infection. Both the recombinant protein rBjRPL27 and the synthetic peptide BjRPL2751-72 effectively killed the Gram-positive bacterium Staphylococcus aureus and the Gram-negative bacterium Aeromonas hydrophila via a combined action of disrupting cell membrane integrity, inducing membrane depolarization, and increasing intracellular reactive oxygen species (ROS) production. Additionally, the sequence of BjRPL2751-72 was highly conserved among all eukaryotic RPL27s, implying an ancient origin for the antibacterial activity of the RPL27 family. In vivo assays showed that BjRPL2751-72 not only efficiently protected zebrafish from A. hydrophila infection, but also killed the bacterium S. aureus on the skin wound of rats. Furthermore, neither BjRPL27 nor BjRPL2751-72 exhibited hemolytic activity towards human red blood cells, making them promising lead molecules for designing novel AMPs. These findings highlight the potential of BjRPL2751-72 as a novel AMP with selective bactericidal properties.

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.

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.

Duchenne muscular dystrophy (DMD), caused by loss-of-function mutations in the dystrophin gene, results in progressive muscle weakness and early fatality. Impaired autophagy is one of the cellular hallmarks of DMD, contributing to the disease progression. Molecular mechanisms underlying the inhibition of autophagy in DMD are not well understood. In the current study, the DMD mouse model mdx was used for the investigation of signaling pathways leading to suppression of autophagy. Mammalian target of rapamycin complex 1 (mTORC1) was hyperactive in the DMD muscles, accompanying muscle weakness and autophagy impairment. Surprisingly, Akt, a well-known upstream regulator of mTORC1, was not responsible for mTORC1 activation or the dystrophic muscle phenotypes. Instead, leucyl-tRNA synthetase (LeuRS) was overexpressed in mdx muscles compared with the wild type. LeuRS activates mTORC1 in a noncanonical mechanism that involves interaction with RagD, an activator of mTORC1. Disrupting LeuRS interaction with RagD by the small-molecule inhibitor BC-LI-0186 reduced mTORC1 activity, restored autophagy, and ameliorated myofiber damage in the mdx muscles. Furthermore, inhibition of LeuRS by BC-LI-0186 improved dystrophic muscle strength in an autophagy-dependent manner. Taken together, our findings uncovered a noncanonical function of the housekeeping protein LeuRS as a potential therapeutic target in the treatment of DMD.

Glutamyl-prolyl-tRNA synthetase (EPRS1) is a bifunctional aminoacyl-tRNA-synthetase (aaRS) essential for decoding the genetic code. EPRS1 resides, with seven other aaRSs and three noncatalytic proteins, in the cytoplasmic multi-tRNA synthetase complex (MSC). Multiple MSC-resident aaRSs, including EPRS1, exhibit stimulus-dependent release from the MSC to perform noncanonical activities distinct from their primary function in protein synthesis. Here, we show EPRS1 is present in both cytoplasm and nucleus of breast cancer cells with constitutively low phosphatase and tensin homolog (PTEN) expression. EPRS1 is primarily cytosolic in PTEN-expressing cells, but chemical or genetic inhibition of PTEN, or chemical or stress-mediated activation of its target, AKT, induces EPRS1 nuclear localization. Likewise, preferential nuclear localization of EPRS1 was observed in invasive ductal carcinoma that were also P-Ser473-AKT+. EPRS1 nuclear transport requires a nuclear localization signal (NLS) within the linker region that joins the catalytic glutamyl-tRNA synthetase and prolyl-tRNA synthetase domains. Nuclear EPRS1 interacts with poly(ADP-ribose) polymerase 1 (PARP1), a DNA-damage sensor that directs poly(ADP-ribosyl)ation (PARylation) of proteins. EPRS1 is a critical regulator of PARP1 activity as shown by markedly reduced ADP-ribosylation in EPRS1 knockdown cells. Moreover, EPRS1 and PARP1 knockdown comparably alter the expression of multiple tumor-related genes, inhibit DNA-damage repair, reduce tumor cell survival, and diminish tumor sphere formation by breast cancer cells. EPRS1-mediated regulation of PARP1 activity provides a mechanistic link between PTEN loss in breast cancer cells, PARP1 activation, and cell survival and tumor growth. Targeting the noncanonical activity of EPRS1, without inhibiting canonical tRNA ligase activity, provides a therapeutic approach potentially supplementing existing PARP1 inhibitors.

Aminoacyl-tRNA synthetases (ARSs) aminoacylate tRNA molecules with their cognate amino acid, enabling information transmission and providing substrates for protein biosynthesis. They also take part in nontranslational functions, mediated by the presence of other proteins domains. Mutations in ARS genes have been described as responsive to numerous factors, including neurological, autoimmune, and oncological. Variants of the ARS genes, both in heterozygosity and homozygosity, have been reported to be responsible for different pathological pictures in humankind. We present the case of a patient referred in infancy for failure to thrive and acquired microcephaly (head circumference: -5 SD). During follow-up we highlighted: dysphagia (which became increasingly severe until it became incompatible with oral feeding, with gastrostomy implantation, resulting in resolution of feeding difficulties), strabismus, hypotonia. NCV (Nerve Conduction Velocity) showed four limbs neuropathy, neurophysiological examination performed at 2 years of age mainly sensory and demyelinating. Exome sequencing (ES) was performed, detecting two novel compound heterozygous variants in the NARS1 gene (OMIM *108410): NM_004539:c.[662 A > G]; [1155dup], p.[(Asn221Ser)]; [(Arg386Thrfs*19)], inherited from mother and father respectively. In this article, we would like to focus on the presence of progressive dysphagia and severe neurodevelopmental disorder, associated with two novel variants in the NARS1 gene.

Antisynthetase syndrome (ASyS) is now a widely recognized entity within the spectrum of idiopathic inflammatory myopathies. Initially described in patients with a triad of myositis, arthritis, and interstitial lung disease (ILD), its presentation can be diverse. Additional common symptoms experienced by patients with ASyS include Raynaud's phenomenon, mechanic's hand, and fever. Although there is a significant overlap with polymyositis and dermatomyositis, the key distinction lies in the presence of antisynthetase antibodies (ASAs). Up to 10 ASAs have been identified to correlate with a presentation of ASyS, each having manifestations that may slightly differ from others. Despite the proposal of three classification criteria to aid diagnosis, the heterogeneous nature of patient presentations poses challenges. ILD confers a significant burden in patients with ASyS, sometimes manifesting in isolation. Notably, ILD is also often the initial presentation of ASyS, requiring pulmonologists to remain vigilant for an accurate diagnosis. This article will comprehensively review the various aspects of ASyS, including disease presentation, diagnosis, management, and clinical course, with a primary focus on its pulmonary manifestations.

Caulerpa lentillifera is rich in polysaccharides, and its polysaccharides show a significant effect in different biological activities including anti-cancer activity. As an edible algae-derived polysaccharide, exploring the role of colon cancer can better develop the application from a dietary therapy perspective. However, more in-depth studies of C. lentillifera polysaccharide on anti-colon cancer activity and mechanism are needed. In this study, we found that Caulerpa lentillifera polysaccharides (CLP) showed potential anti-colon cancer effect on human colon cancer cell HT29 in monolayer (IC50 = 1.954 mg/mL) and spheroid (IC50 = 0.402 mg/mL). Transcriptomics and metabolomics analyses revealed that CLP had an inhibitory effect on HT29 3D spheroid cells by activating aminoacyl-tRNA biosynthesis as well as arginine and proline metabolism pathways. Furthermore, the anti-colon cancer effects of CLP were confirmed through other human colon cancer cell HCT116 and LoVo in monolayer cells (IC50 = 1.890 mg/mL and 1.437 mg/mL, respectively) and 3D spheroid cells (IC50 = 0.344 mg/mL and 0.975 mg/mL, respectively), and three patient-derived organoids with IC50 values of 6.333-8.780 mg/mL. This study provided basic data for the potential application of CLP in adjuvant therapeutic food for colon cancer on multiple levels, while further investigation of detailed mechanism in vivo was still required.

Toxin- and drug-induced tubulointerstitial nephritis (TIN), characterized by interstitial infiltration of immune cells, frequently necessitates dialysis for patients due to irreversible fibrosis. However, agents modulating interstitial immune cells are lacking. Here, we addressed whether the housekeeping enzyme glutamyl-prolyl-transfer RNA synthetase 1 (EPRS1), responsible for attaching glutamic acid and proline to transfer RNA, modulates immune cell activity during TIN and whether its pharmacological inhibition abrogates fibrotic transformation. The immunological feature following TIN induction by means of an adenine-mixed diet was infiltration of EPRS1high T cells, particularly proliferating T and γδ T cells. The proliferation capacity of both CD4+ and CD8+ T cells, along with interleukin-17 production of γδ T cells, was higher in the kidneys of TIN-induced Eprs1+/+ mice than in the kidneys of TIN-induced Eprs1+/- mice. This discrepancy contributed to the fibrotic amelioration observed in kidneys of Eprs1+/- mice. TIN-induced fibrosis was also reduced in Rag1-/- mice adoptively transferred with Eprs1+/- T cells compared to the Rag1-/- mice transferred with Eprs1+/+ T cells. The use of an EPRS1-targeting small molecule inhibitor (bersiporocin) under clinical trials to evaluate its therapeutic potential against idiopathic pulmonary fibrosis alleviated immunofibrotic aggravation in TIN. EPRS1 expression was also observed in human kidney tissues and blood-derived T cells, and high expression was associated with worse patient outcomes. Thus, EPRS1 may emerge as a therapeutic target in toxin- and drug-induced TIN, modulating the proliferation and activity of infiltrated T cells.

Aminoacyl-tRNA synthetases (AARSs) play an indispensable role in the translation of mRNAs into proteins. It has become amply clear that AARSs also have non-canonical or non-translational, yet essential, functions in a myriad of cellular and developmental processes. In this mini-review we discuss the current understanding of the roles of threonyl-tRNA synthetase (TARS) beyond protein synthesis and the underlying mechanisms. The two proteins in eukaryotes - cytoplasmic TARS1 and mitochondrial TARS2 - exert their non-canonical functions in the regulation of gene expression, cell signaling, angiogenesis, inflammatory responses, and tumorigenesis. The TARS proteins utilize a range of biochemical mechanisms, including assembly of a translation initiation complex, unexpected protein-protein interactions that lead to activation or inhibition of intracellular signaling pathways, and cytokine-like signaling through cell surface receptors in inflammation and angiogenesis. It is likely that new functions and novel mechanisms will continue to emerge for these multi-talented proteins.

Aminoacyl-tRNA synthetases (aaRSs) are essential enzymes responsible for linking a transfer RNA (tRNA) with its cognate amino acid present in all the kingdoms of life. Besides their aminoacyl-tRNA synthetase activity, it was described that many of these enzymes can carry out non-canonical functions. They were shown to be involved in important biological processes such as metabolism, immunity, development, angiogenesis and tumorigenesis. In the present work, we provide evidence that tryptophanyl-tRNA synthetase might be involved in a negative feedback loop mitigating the expression of certain interferon-γ-induced genes. Mining the available TCGA and Gtex data, we found that WARS was highly expressed in cutaneous melanoma (SKCM) compared to other cancers and is of good prognosis for this particular cancer type. WARS expression correlates with genes involved in antigen processing and presentation but also transcription factors involved in IFN-γ signaling such as STAT1. In addition, WARS was found in complex with STAT1 in A375 cells treated with IFN-γ. Finally, we showed that knocking down WARS expression during IFN-γ stimulation further increases the expression of GBP2, APOL1, ISG15, HLA-A and IDO1.

Seryl-tRNA synthetases (SerRSs), members of the aminoacyl-tRNA synthetase family, interact with diverse proteins, enabling SerRSs to enhance their role in the translation of the genetic message or to perform alternative functions in cellular processes beyond translation. Atypical archaeal SerRS interacts with arginyl-tRNA synthetase and proteins of the ribosomal P-stalk to optimize translation through tRNA channeling. The complex between yeast SerRS and peroxin Pex21p provides a connection between translation and peroxisome function. The partnership between Arabidopsis SerRS and BEN1 indicates a link between translation and brassinosteroid metabolism and may be relevant in plant stress response mechanisms. In Drosophila, the unusual heterodimeric mitochondrial SerRS coordinates mitochondrial translation and replication via interaction with LON protease. Evolutionarily conserved interactions of yeast and human SerRSs with m3C32 tRNA methyltransferases indicate coordination between tRNA modification and aminoacylation in the cytosol and mitochondria. Human cytosolic SerRS is a cellular hub protein connecting translation to vascular development, angiogenesis, lipogenesis, and telomere maintenance. When translocated to the nucleus, SerRS acts as a master negative regulator of VEGFA gene expression. SerRS alone or in complex with YY1 and SIRT2 competes with activating transcription factors NFκB1 and c-Myc, resulting in balanced VEGFA expression important for proper vascular development and angiogenesis. In hypoxia, SerRS phosphorylation diminishes its binding to the VEGFA promoter, while the lack of nutrients triggers SerRS glycosylation, reducing its nuclear localization. Additionally, SerRS binds telomeric DNA and cooperates with the shelterin protein POT1 to regulate telomere length and cellular senescence. As an antitumor and antiangiogenic factor, human cytosolic SerRS appears to be a promising drug target and therapeutic agent for treating cancer, cardiovascular diseases, and possibly obesity and aging.

Our previous study has reported that supplementation of oligosaccharide-based polymer enhances gut health and disease resistance of pigs infected with enterotoxigenic E. coli (ETEC) F18 in a manner similar to carbadox. The objective of this study was to investigate the impacts of oligosaccharide-based polymer or antibiotic on the host metabolic profiles and colon microbiota of weaned pigs experimentally infected with ETEC F18.

Multivariate analysis highlighted the differences in the metabolic profiles of serum and colon digesta which were predominantly found between pigs supplemented with oligosaccharide-based polymer and antibiotic. The relative abundance of metabolic markers of immune responses and nutrient metabolisms, such as amino acids and carbohydrates, were significantly differentiated between the oligosaccharide-based polymer and antibiotic groups (q < 0.2 and fold change > 2.0). In addition, pigs in antibiotic had a reduced (P < 0.05) relative abundance of Lachnospiraceae and Lactobacillaceae, whereas had greater (P < 0.05) Clostridiaceae and Streptococcaceae in the colon digesta on d 11 post-inoculation (PI) compared with d 5 PI.

The impact of oligosaccharide-based polymer on the metabolic and microbial profiles of pigs is not fully understood, and further exploration is needed. However, current research suggest that various mechanisms are involved in the enhanced disease resistance and performance in ETEC-challenged pigs by supplementing this polymer.

Tire wear particles (TWPs) generated by vehicle tires are ubiquitous in soil ecosystems, while their impact on soil biota remains poorly understood. In this study, we investigated the effects of TWPs (0.1%, 0.7%, and 1.5% of dry soil weight) on the growth and metabolism of mung bean (Vigna radiata) plants over 32 days in soil pots. We found that TWPs-treated soils had high levels of heavy metals and polycyclic aromatic hydrocarbons (PAHs). However, there was no significant impact of TWPs exposure on plant growth, suggesting that mung bean plants have a degree of tolerance to TWPs. Despite the lack of impact on plant growth, exposure to TWPs had significant effects on soil enzyme activities, with a decrease of over 50% in urease and dehydrogenase activity. Furthermore, TWPs exposure resulted in marked changes in the plant metabolite profile, including altered levels of sugars, carboxylic acids, and amino acids, indicating altered nitrogen and amino acid-related metabolic pathways. TWPs exposure also disrupted the rhizospheric and bulk soil microbiota, with a decrease in the abundance of bacterial (Blastococcus) and fungal (Chaetomium) genera involved in nitrogen cycles and suppressing plant diseases. In summary, our study provides new insights into the effects of TWPs on plants and soil, highlighting the potential ecological consequences of TWPs pollution in terrestrial ecosystems and underscoring the need for further research in this area.

Plasmodium is an obligate intracellular parasite that has numerous interactions with different hosts during its elaborate life cycle. This is also the case for the other parasites belonging to the same phylum Apicomplexa. In this study, we bioinformatically identified the components of the multi-synthetase complexes (MSCs) of several Apicomplexa parasites and modelled their assembly using AlphaFold2. It appears that none of these MSCs resemble the two MSCs that we have identified and characterized in Plasmodium. Indeed, tRip, the central protein involved in the association of the two Plasmodium MSCs is different from its homologues, suggesting also that the tRip-dependent import of exogenous tRNAs is not conserved in other apicomplexan parasites. Based on this observation, we searched for obvious differences that could explain the singularity of Plasmodium protein synthesis by comparing tRNA genes and amino acid usage in the different genomes. We noted a contradiction between the large number of asparagine residues used in Plasmodium proteomes and the single gene encoding the tRNA that inserts them into proteins. This observation remains true for all the Plasmodia strains studied, even those that do not contain long asparagine homorepeats.

Aminoacyl-tRNA synthetases (aaRSs) are ancient enzymes that serve a foundational role in the efficient and accurate translation of genetic information from messenger RNA to proteins. These proteins play critical, non-canonical functions in a multitude of cellular processes. Multiple viruses are known to hijack the functions of aaRSs for proviral outcomes, while cells modify antiviral responses through non-canonical functions of certain synthetases. Recent findings have revealed that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the etiological agent of coronaviral disease 19 (COVID-19), utilizes canonical and non-canonical functions of aaRSs, establishing a complex interplay of viral proteins, cellular factors and host aaRSs. In a striking example, an unconventional multi-aaRS complex consisting of glutamyl-prolyl-, lysyl-, arginyl- and methionyl-tRNA synthetases interact with a previously unknown RNA-element in the 3'-end of SARS-CoV-2 genomic and subgenomic RNAs. This review aims to highlight the aaRS-SARS-CoV-2 interactions identified to date, with possible implications for the biology of host aaRSs in SARS-CoV-2 infection.

Human lysyl-tRNA synthetase (LysRS) was previously shown to be re-localized from its normal cytoplasmic location in a multi-aminoacyl-tRNA synthetase complex (MSC) to the nucleus of HIV-1 infected cells. Nuclear localization depends on S207 phosphorylation but the nuclear function of pS207-LysRS in the HIV-1 lifecycle is unknown. Here, we show that HIV-1 replication was severely reduced in a S207A-LysRS knock-in cell line generated by CRISPR/Cas9; this effect was rescued by S207D-LysRS. LysRS phosphorylation up-regulated HIV-1 transcription, as did direct transfection of Ap4A, an upstream transcription factor 2 (USF2) activator that is synthesized by pS207-LysRS. Overexpressing an MSC-derived peptide known to stabilize LysRS MSC binding inhibited HIV-1 replication. Transcription of HIV-1 proviral DNA and other USF2 target genes was reduced in peptide-expressing cells. We propose that nuclear pS207-LysRS generates Ap4A, leading to activation of HIV-1 transcription. Our results suggest a new role for nuclear LysRS in facilitating HIV-1 replication and new avenues for antiviral therapy.

Aminoacyl-tRNA synthetases (aaRSs) are fundamental components of the protein translation machinery. In light of their pivotal role in protein synthesis and structural divergence among species, they have always been considered potential targets for the development of antimicrobial compounds. Arginyl-tRNA synthetase from Trypanosoma cruzi (TcArgRS), the parasite responsible for causing Chagas Disease, contains a 100-amino acid insertion that was found to be completely absent in the human counterpart of similar length, as ascertained from multiple sequence alignment results. Thus, we were prompted to perform a preliminary characterization of TcArgRS using biophysical, biochemical, and bioinformatics tools. We expressed the protein in E. coli and validated its in-vitro enzymatic activity. Additionally, analysis of DTNB kinetics, Circular dichroism (CD) spectra, and ligand-binding studies using intrinsic tryptophan fluorescence measurements aided us to understand some structural features in the absence of available crystal structures. Our study indicates that TcArgRS can discriminate between L-arginine and its analogues. Among the many tested substrates, only L-canavanine and L-thioarginine, a synthetic arginine analogue exhibited notable activation. The binding of various substrates was also determined using in silico methods. This study may provide a viable foundation for studying small compounds that can be targeted against TcArgRS.

Neurons build synaptic contacts using different protein combinations that define the specificity, function, and plasticity potential of synapses; however, the diversity of synaptic proteomes remains largely unexplored. We prepared synaptosomes from 7 different transgenic mouse lines with fluorescently labeled presynaptic terminals. Combining microdissection of 5 different brain regions with fluorescent-activated synaptosome sorting (FASS), we isolated and analyzed the proteomes of 18 different synapse types. We discovered ∼1,800 unique synapse-type-enriched proteins and allocated thousands of proteins to different types of synapses (https://syndive.org/). We identify shared synaptic protein modules and highlight the proteomic hotspots for synapse specialization. We reveal unique and common features of the striatal dopaminergic proteome and discover the proteome signatures that relate to the functional properties of different interneuron classes. This study provides a molecular systems-biology analysis of synapses and a framework to integrate proteomic information for synapse subtypes of interest with cellular or circuit-level experiments.

The resistance of microorganisms to antimicrobials has endangered the health of many people across the world. Overcoming the resistance problem will require the invention of molecules with a new mechanism of action so that no cross-resistance with existing therapies occurs. Because of their powerful antibacterial activity against a wide spectrum of Gram-positive and Gram-negative bacterial strains, heterocyclic compounds are appealing candidates for medicinal chemists. In this regard, as unique hybrid compounds, we synthesized a novel family of bis-thiazoles linked to quinoxaline or thienothiophene via the 2-phenoxy-N-arylacetamide moiety. The target compounds were synthesized by reacting the relevant bis(α-haloketones) with the corresponding thiosemicarbazones in EtOH at reflux with a few drops of TEA. Under comparable reaction conditions, the isomeric bis(thiazoles) were synthesized by reacting the appropriate bis(thiosemicarbazone) with the respective α-haloketones. The structures of the novel compounds were confirmed using elements and spectral data. All of the synthesized compounds were tested for antibacterial activity in vitro. With an inhibitory zone width of 12 mm, compound 12a had the same activity as the reference medication tobramycin against Staphylococcus aureus. Compound 12b showed 20 mg/mL as a minimum inhibitory concentration (MIC) against Bacillus subtilis. Some of the synthesized compounds were tested via molecular docking against two bacterial proteins (dihydrofolate reductase and tyrosyl-tRNA synthetase).

Transfer RNAs (tRNAs) have been historically viewed as non-dynamic adaptors that decode the genetic code into proteins. Recent work has uncovered dynamic regulatory roles for these fascinating molecules. Advances in tRNA detection methods have revealed that specific tRNAs can become modulated upon DNA copy number and chromatin alterations and can also be perturbed by oncogenic signalling and transcriptional regulators in cancer cells or the tumour microenvironment. Such alterations in the levels of specific tRNAs have been shown to causally impact cancer progression, including metastasis. Moreover, sequencing methods have identified tRNA-derived small RNAs that influence various aspects of cancer progression, such as cell proliferation and invasion, and could serve as diagnostic and prognostic biomarkers or putative therapeutic targets in various cancers. Finally, there is accumulating evidence, including from genetic models, that specific tRNA synthetases - the enzymes responsible for charging tRNAs with amino acids - can either promote or suppress tumour formation. In this Review, we provide an overview of how deregulation of tRNAs influences cancer formation and progression.

Previous studies have shown that the elevated levels of circulating branched-chain amino acids (BCAAs) are associated with the development of insulin resistance and its complications, including obesity, type 2 diabetes, cardiovascular disease and some cancers. However, animal models that can mimic the metabolic state of chronically elevated BCAAs in humans are rare. Therefore, the aim of this study was to establish the above animal model and analyse the metabolic changes associated with high BCAA levels. Sixteen 8-week-old Sprague-Dawley (SD) rats were randomly divided into two groups and given either a high fructose diet or a normal diet. BCAA levels as well as blood glucose and lipid levels were measured at different time points of feeding. The mRNA expression levels of two key enzymes of BCAA catabolism, ACAD (acyl-CoA dehydrogenase) and BCKDH (branched-chain α-keto acid dehydrogenase), were measured by qPCR, and the protein expression levels of these two enzymes were analysed by immunohistochemistry. Finally, the metabolite expression differences between the two groups were analysed by Q300 metabolomics technology. Our study confirms that defects in the catabolic pathways of BCAAs lead to increased levels of circulating BCAAs, resulting in disorders of glucose and lipid metabolism characterized by insulin resistance by affecting metabolic pathways associated with amino acids and bile acids.

Efficient communication between mitochondria and the nucleus underlies homoeostatic metabolic control, though the involved mitochondrial factors and their mechanisms are poorly defined. Here, we report the surprising detection of multiple mitochondrial-derived transfer RNAs (mito-tRNAs) within the nuclei of human cells. Focused studies of nuclear-transported mito-tRNA-asparagine (mtAsn) revealed that its cognate charging enzyme (NARS2) is also present in the nucleus. MtAsn promoted interaction of NARS2 with histone deacetylase 2 (HDAC2), and repressed HDAC2 association with specific chromatin loci. Perturbation of this axis using antisense oligonucleotides promoted nucleotide biogenesis and enhanced breast cancer growth, and RNA and nascent transcript sequencing demonstrated specific alterations in the transcription of nuclear genes. These findings uncover nucleic-acid mediated communication between two organelles and the existence of a machinery for nuclear gene regulation by a mito-tRNA that restricts tumor growth through metabolic control.

Multiple mitochondrial-derived tRNAs are detected in human cell nucleiMtAsn promotes binding between NARS2 and HDAC2Metabolic alterations driven by mtAsn impact cell proliferationMtAsn inhibition releases HDAC2 to bind and transcriptionally regulate multiple nuclear genes.

Recent studies demonstrate that extracellular-released aminoacyl-tRNA synthetases (aaRSs) play unique roles in immune responses and diseases. This study aimed to understand the role of extracellular aaRSs in the pathogenesis of rheumatoid arthritis (RA).

Primary macrophages and fibroblast-like synoviocytes were cultured with aaRSs. aaRS-induced cytokine production including IL-6 and TNF-α was detected by ELISA. Transcriptomic features of aaRS-stimulated macrophages were examined using RNA-sequencing. Serum and synovial fluid (SF) aaRS levels in patients with RA were assessed using ELISA. Peptidyl arginine deiminase (PAD) 4 release from macrophages stimulated with aaRSs was detected by ELISA. Citrullination of aaRSs by themselves was examined by immunoprecipitation and western blotting. Furthermore, aaRS inhibitory peptides were used for inhibition of arthritis in two mouse RA models, collagen-induced arthritis and collagen antibody-induced arthritis.

All 20 aaRSs functioned as alarmin; they induced pro-inflammatory cytokines through the CD14-MD2-TLR4 axis. Stimulation of macrophages with aaRSs displayed persistent innate inflammatory responses. Serum and SF levels of many aaRSs increased in patients with RA compared with control subjects. Furthermore, aaRSs released PAD4 from living macrophages, leading to their citrullination. We demonstrate that aaRS inhibitory peptides suppress cytokine production and PAD4 release by aaRSs and alleviate arthritic symptoms in a mouse RA model.

Our findings uncovered the significant role of aaRSs as a novel alarmin in RA pathogenesis, indicating that their blocking agents are potent antirheumatic drugs.

Aminoacyl-tRNA synthetases form the protein family that controls the interpretation of the genetic code, with tRNA aminoacylation being the key chemical step during which an amino acid is assigned to a corresponding sequence of nucleic acids. In consequence, aminoacyl-tRNA synthetases have been studied in their physiological context, in disease states, and as tools for synthetic biology to enable the expansion of the genetic code. Here, we review the fundamentals of aminoacyl-tRNA synthetase biology and classification, with a focus on mammalian cytoplasmic enzymes. We compile evidence that the localization of aminoacyl-tRNA synthetases can be critical in health and disease. In addition, we discuss evidence from synthetic biology which made use of the importance of subcellular localization for efficient manipulation of the protein synthesis machinery. This article is categorized under: RNA Processing Translation > Translation Regulation RNA Processing > tRNA Processing RNA Export and Localization > RNA Localization.

The function of Seryl-tRNA synthetase in fungi during gene transcription regulation beyond translation has not been reported. Here, we report a seryl-tRNA synthetase, ThserRS, which can negatively regulate laccase lacA transcription in Trametes hirsuta AH28-2 under exposure to copper ion. ThserRS was obtained through yeast one-hybrid screening using a bait sequence of lacA promoter (-502 to -372 bp). ThserRS decreased while lacA increased at the transcription level in T. hirsuta AH28-2 in the first 36 h upon CuSO4 induction. Then, ThserRS was upregulated, and lacA was downregulated. ThserRS overexpression in T. hirsuta AH28-2 resulted in a decrement in lacA transcription and LacA activity. By comparison, ThserRS silencing led to increased LacA transcripts and activity. A minimum of a 32-bp DNA fragment containing two putative xenobiotic response elements could interact with ThserRS, with a dissociation constant of 919.9 nM. ThserRS localized in the cell cytoplasm and nucleus in T. hirsuta AH28-2 and was heterologously expressed in yeast. ThserRS overexpression also enhanced mycelial growth and oxidative stress resistance. The transcriptional level of several intracellular antioxidative enzymes in T. hirsuta AH28-2 was upregulated. Our results demonstrate a noncanonical activity of SerRS that acts as a transcriptional regulation factor to upregulate laccase expression at an early stage after exposure to copper ions. IMPORTANCE Seryl-tRNA synthetase is well known for the attachment of serine to the corresponding cognate tRNA during protein translation. In contrast, its functions beyond translation in microorganisms are underexplored. We performed in vitro and cell experiments to show that the seryl-tRNA synthetase in fungi with no UNE-S domain at the carboxyl terminus can enter the nucleus, directly interact with the promoter of the laccase gene, and negatively regulate the fungal laccase transcription early upon copper ion induction. Our study deepens our understanding of the Seryl-tRNA synthetase noncanonical activities in microorganisms. It also demonstrates a new transcription factor for fungal laccase transcription.

There has been an increase in the number of women suffering from breast cancer in recent years, and discovering new therapeutic targets and efficacy predictive markers is critical for comprehensive breast cancer treatment.

First, we used bioinformatics methods to analyze TARS1(encoding cytoplasmicthreonyl-tRNA synthetase) expression, prognosis, and clinicopathological characteristics in TCGA database breast cancers, and then we collected breast cancer specimens from our center for validation. TARS1 was then subjected to GSEA (Gene Set Enrichment Analysis) enrichment analysis, GO/KEGG pathway enrichment analysis, and breast cancer immune infiltration characterization. As a last step, we examined TARS1's effects on breast cancer cell behavior with cellular assays.

The overexpression of TARS1 has been found in several malignant tumors, including breast cancer, and has been linked to poor prognoses. Breast cancers with large primary tumors and negative hormone receptors are more likely to overexpress TARS1. Overexpression of TARS1 promotes the infiltration of T cells, such as Tregs and Th2s, while inhibiting the infiltration of NK cells and CD8+ T cells, which are anticancer cells in breast cancer. TARS1 was also found to be co-expressed with the majority of immune checkpoint-related genes, and breast cancer with TARS1 overexpression responded better to immunotherapy. By knocking down TARS1, breast cancer cells were prevented from proliferating and invading, as well as exhibiting other malignant biological properties.

According to our study, TARS1 may be an oncogene in breast cancer and may be a biomarker of efficacy or a target of immunotherapy in breast cancer.

Various stress conditions are signaled through phosphorylation of translation initiation factor eukaryotic initiation factor 2α (eIF2α) to inhibit global translation while selectively activating transcription factor ATF4 to aid cell survival and recovery. However, this integrated stress response is acute and cannot resolve lasting stress. Here, we report that tyrosyl-tRNA synthetase (TyrRS), a member of the aminoacyl-tRNA synthetase family that responds to diverse stress conditions through cytosol-nucleus translocation to activate stress-response genes, also inhibits global translation. However, it occurs at a later stage than eIF2α/ATF4 and mammalian target of rapamycin (mTOR) responses. Excluding TyrRS from the nucleus over-activates translation and increases apoptosis in cells under prolonged oxidative stress. Nuclear TyrRS transcriptionally represses translation genes by recruiting TRIM28 and/or NuRD complex. We propose that TyrRS, possibly along with other family members, can sense a variety of stress signals through intrinsic properties of this enzyme and strategically located nuclear localization signal and integrate them by nucleus translocation to effect protective responses against chronic stress.

Commensal bacteria are critically involved in the establishment of tolerance against inflammatory challenges, the molecular mechanisms of which are just being uncovered. All kingdoms of life produce aminoacyl-tRNA synthetases (ARSs). Thus far, the non-translational roles of ARSs have largely been reported in eukaryotes. Here, we report that the threonyl-tRNA synthetase (AmTARS) of the gut-associated bacterium Akkermansia muciniphila is secreted and functions to monitor and modulate immune homeostasis. Secreted AmTARS triggers M2 macrophage polarization and orchestrates the production of anti-inflammatory IL-10 via its unique, evolutionary-acquired regions, which mediates specific interactions with TLR2. This interaction activates the MAPK and PI3K/AKT signaling pathways, which converge on CREB, leading to an efficient production of IL-10 and suppression of the central inflammatory mediator NF-κB. AmTARS restores IL-10-positive macrophages, increases IL-10 levels in the serum, and attenuates the pathological effects in colitis mice. Thus, commensal tRNA synthetases can act as intrinsic mediators that maintain homeostasis.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of COVID-19, generates multiple protein-coding, subgenomic RNAs (sgRNAs) from a longer genomic RNA, all bearing identical termini with poorly understood roles in regulating viral gene expression. Insulin and interferon-gamma, two host-derived, stress-related agents, and virus spike protein, induce binding of glutamyl-prolyl-tRNA synthetase (EPRS1), within an unconventional, tetra-aminoacyl-tRNA synthetase complex, to the sgRNA 3'-end thereby enhancing sgRNA expression. We identify an EPRS1-binding sarbecoviral pan-end activating RNA (SPEAR) element in the 3'-end of viral RNAs driving agonist-induction. Translation of another co-terminal 3'-end feature, ORF10, is necessary for SPEAR-mediated induction, independent of Orf10 protein expression. The SPEAR element enhances viral programmed ribosomal frameshifting, thereby expanding its functionality. By co-opting noncanonical activities of a family of essential host proteins, the virus establishes a post-transcriptional regulon stimulating global viral RNA translation. A SPEAR-targeting strategy markedly reduces SARS-CoV-2 titer, suggesting a pan-sarbecoviral therapeutic modality.

Monoclonal antibody (mAb)-based biologics are well established treatments of cancer. Antibody discovery campaigns are typically directed at a single target of interest, which inherently limits the possibility of uncovering novel antibody specificities or functionalities. Here, we present a target-unbiased approach for antibody discovery that relies on generating mAbs against native target cell surfaces via phage display. This method combines a previously reported method for improved whole-cell phage display selections with next-generation sequencing analysis to efficiently identify mAbs with the desired target cell reactivity. Applying this method to multiple myeloma cells yielded a panel of >50 mAbs with unique sequences and diverse reactivities. To uncover the identities of the cognate antigens recognized by this panel, representative mAbs from each unique reactivity cluster were used in a multi-omic target deconvolution approach. From this, we identified and validated three cell surface antigens: PTPRG, ICAM1, and CADM1. PTPRG and CADM1 remain largely unstudied in the context of multiple myeloma, which could warrant further investigation into their potential as therapeutic targets. These results highlight the utility of optimized whole-cell phage display selection methods and could motivate further interest in target-unbiased antibody discovery workflows.

Aminoacyl-tRNA synthetases (ARSs) play an essential role in protein synthesis, being responsible for ligating tRNA molecules to their corresponding amino acids in a reaction known as 'tRNA aminoacylation'. Separate ARSs carry out the aminoacylation reaction in the cytosol and in mitochondria, and mutations in almost all ARS genes cause pathophysiology most evident in the nervous system. Dominant mutations in multiple cytosolic ARSs have been linked to forms of peripheral neuropathy including Charcot-Marie-Tooth disease, distal hereditary motor neuropathy, and spinal muscular atrophy. This review provides an overview of approaches that have been employed to model each of these diseases in vivo, followed by a discussion of the existing animal models of dominant ARS disorders and key mechanistic insights that they have provided. In summary, ARS disease models have demonstrated that loss of canonical ARS function alone cannot fully account for the observed disease phenotypes, and that pathogenic ARS variants cause developmental defects within the peripheral nervous system, despite a typically later onset of disease in humans. In addition, aberrant interactions between mutant ARSs and other proteins have been shown to contribute to the disease phenotypes. These findings provide a strong foundation for future research into this group of diseases, providing methodological guidance for studies on ARS disorders that currently lack in vivo models, as well as identifying candidate therapeutic targets.

Protein synthesis is a fundamental process that underpins almost every aspect of cellular functioning. Intriguingly, despite their common function, recessive mutations in aminoacyl-tRNA synthetases (ARSs), the family of enzymes that pair tRNA molecules with amino acids prior to translation on the ribosome, cause a diverse range of multi-system disorders that affect specific groups of tissues. Neurological development is impaired in most ARS-associated disorders. In addition to central nervous system defects, diseases caused by recessive mutations in cytosolic ARSs commonly affect the liver and lungs. Patients with biallelic mutations in mitochondrial ARSs often present with encephalopathies, with variable involvement of peripheral systems. Many of these disorders cause severe disability, and as understanding of their pathogenesis is currently limited, there are no effective treatments available. To address this, accurate in vivo models for most of the recessive ARS diseases are urgently needed. Here, we discuss approaches that have been taken to model recessive ARS diseases in vivo, highlighting some of the challenges that have arisen in this process, as well as key results obtained from these models. Further development and refinement of animal models is essential to facilitate a better understanding of the pathophysiology underlying recessive ARS diseases, and ultimately to enable development and testing of effective therapies.

Aminoacyl-tRNA synthetases (aaRSs) are indispensable players in translation. Usually, two or three genes encode cytoplasmic and mitochondrial threonyl-tRNA synthetases (ThrRSs) in eukaryotes. Here, we reported that Caenorhabditis elegans harbors only one tars-1, generating cytoplasmic and mitochondrial ThrRSs via translational reinitiation. Mitochondrial tars-1 knockdown decreased mitochondrial tRNA^Thr charging and translation and caused pleotropic phenotypes of delayed development, decreased motor ability and prolonged lifespan, which could be rescued by replenishing mitochondrial tars-1. Mitochondrial tars-1 deficiency leads to compromised mitochondrial functions including the decrease in oxygen consumption rate, complex Ⅰ activity and the activation of the mitochondrial unfolded protein response (UPR^mt), which contributes to longevity. Furthermore, deficiency of other eight mitochondrial aaRSs in C. elegans and five in mammal also caused activation of the UPR^mt. In summary, we deciphered the mechanism of one tars-1, generating two aaRSs, and elucidated the biochemical features and physiological function of C. elegans tars-1. We further uncovered a conserved connection between mitochondrial translation deficiency and UPR^mt.

Dinucleoside polyphosphates, a class of nucleotides found amongst all the Trees of Life, have been gathering a lot of attention in the past decades due to their putative role as cellular alarmones. In particular, diadenosine tetraphosphate (AP4A) has been widely studied in bacteria facing various environmental challenges and has been proposed to be important for ensuring cellular survivability through harsh conditions. Here, we discuss the current understanding of AP4A synthesis and degradation, protein targets, their molecular structure where possible, and insights into the molecular mechanisms of AP4A action and its physiological consequences. Lastly, we will briefly touch on what is known with regards to AP4A beyond the bacterial kingdom, given its increasing appearance in the eukaryotic world. Altogether, the notion that AP4A is a conserved second messenger in organisms ranging from bacteria to humans and is able to signal and modulate cellular stress regulation seems promising.

Aminoacyl-tRNA synthetases (aaRSs) are essential components for mRNA translation. Two sets of aaRSs are required for cytoplasmic and mitochondrial translation in vertebrates. Interestingly, TARSL2 is a recently evolved duplicated gene of TARS1 (encoding cytoplasmic threonyl-tRNA synthetase) and represents the only duplicated aaRS gene in vertebrates. Although TARSL2 retains the canonical aminoacylation and editing activities in vitro, whether it is a true tRNA synthetase for mRNA translation in vivo is unclear. In this study, we showed that Tars1 is an essential gene since homozygous Tars1 knockout mice were lethal. In contrast, when Tarsl2 was deleted in mice and zebrafish, neither the abundance nor the charging levels of tRNA^Thrs were changed, indicating that cells relied on Tars1 but not on Tarsl2 for mRNA translation. Furthermore, Tarsl2 deletion did not influence the integrity of the multiple tRNA synthetase complex (MSC), suggesting that Tarsl2 is a peripheral member of the MSC. Finally, we observed that Tarsl2-deleted mice exhibited severe developmental retardation, elevated metabolic capacity, and abnormal bone and muscle development after 3 weeks. Collectively, these data suggest that, despite its intrinsic activity, loss of Tarsl2 has little influence on protein synthesis but does affect mouse development.