tRip is a tRNA import protein specific to Plasmodium, the causative agent of malaria. In addition to its membrane localization and tRNA trafficking properties, tRip has the capacity to associate with ...three aminoacyl‐tRNA synthetases (aaRS), the glutamyl‐ (ERS), glutaminyl‐ (QRS), and methionyl‐ (MRS) tRNA synthetases. In eukaryotes, such multi‐aaRSs complexes (MSC) regulate the moonlighting activities of aaRSs. In Plasmodium, tRip and the three aaRSs all contain an N‐terminal GST‐like domain involved in the assembly of two independent complexes: the Q‐complex (tRip:ERS:QRS) and the M‐complex (tRip:ERS:MRS) with a 2:2:2 stoichiometry and in which the association of the GST‐like domains of tRip and ERS (tRip‐N:ERS‐N) is central. In this study, the crystal structure of the N‐terminal GST‐like domain of ERS was solved and made possible further investigation of the solution architecture of the Q‐ and M‐complexes by small‐angle x‐ray scattering (SAXS). This strategy relied on the engineering of a tRip‐N‐ERS‐N chimeric protein to study the structural scaffold of both Plasmodium MSCs and confirm the unique homodimerization pattern of tRip in solution. The biological impact of these structural arrangements is discussed.
PDB Code(s): 8BCQ and 8BHD
Abstract
The number of tRNAs encoded in plant mitochondrial genomes varies considerably. Ongoing loss of bacterial-like mitochondrial tRNA genes in many lineages necessitates the import of ...nuclear-encoded counterparts that share little sequence similarity. Because tRNAs are involved in highly specific molecular interactions, this replacement process raises questions about the identity and trafficking of enzymes necessary for the maturation and function of newly imported tRNAs. In particular, the aminoacyl-tRNA synthetases (aaRSs) that charge tRNAs are usually divided into distinct classes that specialize on either organellar (mitochondrial and plastid) or nuclear-encoded (cytosolic) tRNAs. Here, we investigate the evolution of aaRS subcellular localization in a plant lineage (Sileneae) that has experienced extensive and rapid mitochondrial tRNA loss. By analyzing full-length mRNA transcripts (PacBio Iso-Seq), we found predicted retargeting of many ancestrally cytosolic aaRSs to the mitochondrion and confirmed these results with colocalization microscopy assays. However, we also found cases where aaRS localization does not appear to change despite functional tRNA replacement, suggesting evolution of novel interactions and charging relationships. Therefore, the history of repeated tRNA replacement in Sileneae mitochondria reveals that differing constraints on tRNA/aaRS interactions may determine which of these alternative coevolutionary paths is used to maintain organellar translation in plant cells.
TriTryp diseases (Leishmaniasis, Human African Trypanosomiasis (HAT), and Chagas disease) are devastating parasitic neglected tropical diseases (NTDs) that affect billions of people in developing ...countries, cause high mortality in humans, and impose a large socio-economic burden. The current treatment options against tritryp diseases are suboptimal and challenging due to the emergence of resistance against available tritryp drugs. Hence, designing and developing effective anti-tritryp drugs with novel targets are required. Aminoacyl-tRNA synthetases (AARSs) involved in specific aminoacylation of transfer RNAs (tRNAs), interrupt protein synthesis through inhibitors, and retard the parasite growth. AaRSs have long been studied as therapeutic targets in bacteria, and three aaRS inhibitors, mupirocin (against IleRS), tavaborole AN2690 (against LeuRS), and halofuginone (against ProRS), are already in clinical practice. The structural differences between tritryp and human aaRSs and the presence of unique sequences (N-terminal domain/C-terminal domain/catalytic domain) make them potential target for developing selective inhibitors. Drugs based on a single aaRS target developed by high-throughput screening (HTS) are less effective due to the emergence of resistance. However, designing multi-targeted drugs may be a better strategy for resistance development. In this perspective, we discuss the characteristics of tritryp aaRSs, sequence conservation in their orthologs and their peculiarities, recent advancements towards the single-target and multi-target aaRS inhibitors developed through rational design.
•Tritryp aminoacyl-tRNA synthetases (aaRSs) are essential for growth and viability of tritryp parasites.•Tritryp aaRS are potential drug target due to presence of unique sequence peculiarities and structural difference between tritryp and human aaRSs.•Multi-targeted drug may be better strategy for developing less resistant and more effective drug against trirtyp.•Recently identified tritryp specific insertions/extensions or domains may be targeted for the development of selective tritryp aaRS inhibitors.
Aminoacyl-tRNA synthetases (ARSs) are enzymes that ligate their cognate amino acids to tRNAs for protein synthesis. However, recent studies have shown that their functions are expanded beyond protein ...synthesis through the interactions with diverse cellular factors. In this review, we discuss how ARSs have evolved to expand and control their functions by forming protein assemblies. We particularly focus on a macromolecular ARS complex in eukaryotes, named multi-tRNA synthetase complex (MSC), which is proposed to provide a channel through which tRNAs reach bound ARSs to receive their cognate amino acid and transit further to the translation machinery. Approximately half of the ARSs assemble into the MSC through cis-acting noncatalytic domains attached to their catalytic domains and trans-acting factors. Evolution of the MSC included its functional expansion, during which the MSC interaction network was augmented by additional cellular pathways present in higher eukaryotes. We also discuss MSC components that could be functionally involved in the pathophysiology of tumorigenesis. For example, the activities of some trans-acting factors have tumor-suppressing effects or maintain DNA integrity and are functionally compromised in cancer. On the basis of Gene Ontology analyses, we propose that the regulatory activities of the MSC-associated ARSs mainly converge on five biological processes, including mammalian target of rapamycin (mTOR) and DNA repair pathways. Future studies are needed to investigate how the MSC-associated and free-ARSs interact with each other and other factors in the control of multiple cellular pathways, and how aberrant or disrupted interactions in the MSC can cause disease.
We have previously identified a unique disulfide bond in the crystal structure of Arabidopsis cytosolic seryl‐tRNA synthetase involving cysteines evolutionarily conserved in all green plants. Here, ...we discovered that both cysteines are important for protein stability, but with opposite effects, and that their microenvironment may promote disulfide bond formation in oxidizing conditions. The crystal structure of the C244S mutant exhibited higher rigidity and an extensive network of noncovalent interactions correlating with its higher thermal stability. The activity of the wild‐type showed resistance to oxidation with H2O2, while the activities of cysteine‐to‐serine mutants were impaired, indicating that the disulfide link may enable the protein to function under oxidative stress conditions which can be beneficial for an efficient plant stress response.
We have examined the role of the disulfide link between evolutionarily conserved cysteines in plant cytosolic seryl‐tRNA synthetase. We have identified features of the protein microenvironment which may promote disulfide bond formation in oxidizing conditions. Activity assays showed that the disulfide link is important for protein resistance to oxidation, which may be beneficial for translation during oxidative stress conditions in plants.
Homochirality of biomacromolecules is a prerequisite for their proper functioning and hence essential for all life forms. This underscores the role of cellular chiral checkpoints in enforcing ...homochirality during protein biosynthesis. d‐Aminoacyl‐tRNA deacylase (DTD) is an enzyme that performs ‘chirality‐based proofreading’ to remove d‐amino acids mistakenly attached to tRNAs, thus recycling them for further rounds of translation. Paradoxically, owing to its l‐chiral rejection mode of action, DTD can remove glycine as well, which is an achiral amino acid. However, this activity is modulated by discriminator base (N73) in tRNA, a unique element that protects the cognate Gly‐tRNAGly. Here, we review our recent work showing various aspects of DTD and tRNAGly coevolution and its key role in maintaining proper translation surveillance in both bacteria and eukaryotes. Moreover, we also discuss two major optimization events on DTD and tRNA that resolved compatibility issues among the archaeal and the bacterial translation apparatuses. Importantly, such optimizations are necessary for the emergence of mitochondria and successful eukaryogenesis.
Here, we summarize various aspects of DTD and tRNAGly co‐evolution and their key role in maintaining proper translation surveillance in both bacteria and eukaryotes. We also discuss two major optimization events on DTD and tRNA resulting from incompatibility issues among the archaeal and bacterial translation apparatuses for the emergence of mitochondria and successful eukaryogenesis.
Mullen et al. generate new models of pathogenic, dominant HARS1 mutations and show that they increase eIF2α phosphorylation and decrease protein translation in neurons. These results are consistent ...with a dominant‐negative mechanism of ARS‐mediated peripheral neuropathy. Comment on: https://doi.org/10.1111/febs.15449
The pathogenic mechanism of neuropathy‐associated aminoacyl‐tRNA synthetase (ARS) gene variants is poorly defined. Mullen et al. generate new models of pathogenic, dominant HARS1 mutations and show that they increase eIF2α phosphorylation and decrease protein translation in neurons. These results are consistent with a dominant‐negative mechanism of ARS‐mediated peripheral neuropathy.
Comment on: https://doi.org/10.1111/febs.15449
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