Human tRNA synthetase catalytic nulls with diverse functions

Lo, Woo-Kuen; Gardiner, Elisabeth; Xu, Zhiwen; Lau, Ching-Fun; Wang, Feng; Zhou, Jie; Mendlein, John; Nangle, Leslie A.; Chiang, Kyle P.; Yang, Xiang‐Lei; Au, K.; Wong, Wing Hung; Guo, Min; Zhang, Mingjie; Schimmel, Paul

doi:10.1126/science.1252943

Cited by 106 publications

(116 citation statements)

References 34 publications

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“…The variant is readily translated and imported into mitochondria, suggesting a noncanonical mitochondrial function. This phenomenon is reminiscent of other examples of synthetase fragments and supports the hypothesis that alternative splicing of mRNA transcripts of aaRS is a way to enlarge aaRS additional functions in higher eukaryotes (27,28). Altogether, our results significantly improve our understanding of hmtThrRS and provide a basis for further investigations of TARS2-related human diseases.…”

supporting

confidence: 61%

“…Existence of an Alternative Splicing Variant with Possible Non-translational Function-Recently, it was reported that nearly all aaRS genes in humans encode multiple splicing variants, creating a large number of aaRS-related catalytic nulls (28). These catalytic nulls usually keep their non-catalytic domains and exhibit deletions or truncations in the catalytic core, inducing loss of tRNA aminoacylation activity.…”

Section: Human Mitochondrial Thrrs Misactivates Ser and Mainly Uses Tmentioning

confidence: 99%

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A Human Disease-causing Point Mutation in Mitochondrial Threonyl-tRNA Synthetase Induces Both Structural and Functional Defects

Wang

Zhou

Ruan

et al. 2016

Journal of Biological Chemistry

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Mitochondria require all translational components, including aminoacyl-tRNA synthetases (aaRSs), to complete organelle protein synthesis. Some aaRS mutations cause mitochondrial disorders, including human mitochondrial threonyl-tRNA synthetase (hmtThrRS) (encoded by TARS2), the P282L mutation of which causes mitochondrial encephalomyopathies. However, its catalytic and structural consequences remain unclear. Herein, we cloned TARS2 and purified the wild-type and P282L mutant hmtThrRS. hmtThrRS misactivates non-cognate Ser and uses post-transfer editing to clear erroneously synthesized products. In vitro and in vivo analyses revealed that the mutation induces a decrease in Thr activation, aminoacylation, and proofreading activities and a change in the protein structure and/or stability, which might cause reduced catalytic efficiency. We also identified a splicing variant of TARS2 mRNA lacking exons 8 and 9, the protein product of which is targeted into mitochondria. In HEK293T cells, the variant does not dimerize and cannot complement the ThrRS knock-out strain in yeast, suggesting that the truncated protein is inactive and might have a noncanonical function, as observed for other aaRS fragments. The present study describes the aminoacylation and editing properties of hmtThrRS, clarifies the molecular consequences of the P282L mutation, and shows that the yeast ThrRS-deletion model is suitable to test pathology-associated point mutations or alternative splicing variants of mammalian aaRS mRNAs.Aminoacyl-tRNA synthetases (aaRSs) 4 supply the ribosome with aminoacyl-tRNA substrates for protein synthesis (1, 2). This process starts with amino acid activation by condensation with ATP to form the aminoacyl adenylate aa-AMP and pyrophosphate. The activated amino acid then reacts with the 3Ј-end of the cognate tRNA to yield the aminoacyl-tRNA (aatRNA), which is transferred by EF-Tu to the protein biosynthesis machinery as a building block. Highly accurate protein synthesis is indispensable for cell growth. To maintain fidelity during protein synthesis, aaRS needs to select the correct amino acid and tRNA substrates from a large number of structurally similar molecules in the cells. The specificity of aaRS is greatly challenged by the presence of the 20 amino acids in which the physicochemical properties are sometimes very similar and by non-proteogenous amino acids and diverse metabolites produced by cell metabolism. aaRSs that do not show an overall selectivity above 1 in 3000 are predicted to require proofreading (editing) activity to maintain sufficient accuracy during aa-tRNA synthesis (3). In fact, editing activity has evolved in half of the currently identified aaRS to remove aberrantly produced aa-AMP (pretransfer editing) and/or aa-tRNA (posttransfer editing) (4). tRNA mischarging may be toxic and deleterious for cells. Even a slight decrease in aminoacylation accuracy could cause an intracellular accumulation of misfolded proteins and up-regulation of cytoplasmic protein chaperones in neurons, leading to severe...

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supporting

confidence: 61%

Section: Human Mitochondrial Thrrs Misactivates Ser and Mainly Uses Tmentioning

confidence: 99%

A Human Disease-causing Point Mutation in Mitochondrial Threonyl-tRNA Synthetase Induces Both Structural and Functional Defects

Wang

Zhou

Ruan

et al. 2016

Journal of Biological Chemistry

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“…Some of these catalytically inactive splice variants fall into the SEP regime in terms of their small size. Interestingly, each of the AARS variants demonstrated novel regulatory activities spanning a variety of physiological processes, including cell differentiation and proliferation, transcriptional regulation, and inflammatory response, which are distinct from the original aminoacylation function (Lo et al, 2014). In addition to the fact that alternative splicing is able to produce functional SEPs, SEPs can also be generated via polycistronic translation of a given RNA transcript.…”

Section: Sep Characteristicsmentioning

confidence: 99%

Identification and characterization of sORF-encoded polypeptides

Chu

Saghatelian

2015

Critical Reviews in Biochemistry and Molecular Biology

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Molecular biology, genomics and proteomics methods have been utilized to reveal a non-annotated class of endogenous polypeptides (small proteins and peptides) encoded by short open reading frames (sORFs), or small open reading frames (smORFs). We refer to these polypeptides as s(m)ORF-encoded polypeptides or SEPs. The early SEPs were identified via genetic screens, and many of the RNAs that contain s(m)ORFs were originally considered to be non-coding; however, elegant work in bacteria and flies demonstrated that these s(m)ORFs code for functional polypeptides as small as 11-amino acids in length. The discovery of these initial SEPs led to searches for these molecules using methods such as ribosome profiling and proteomics, which have revealed the existence of many SEPs, including novel human SEPs. Unlike screens, –omics methods do not necessarily link a SEP to a cellular or biological function, but functional genomic and proteomic strategies have demonstrated that at least some of these newly discovered SEPs have biochemical and cellular functions. Here, we provide an overview of these results and discuss the future directions in this emerging field.

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“…Human TyrRS has been proposed to generate MiniTyrRS and EMAPII fragments by two different mechanisms: proteolytic cleavage 25 and alternative splicing 35 . To test the proteolytic hypothesis, we used transgenic flies expressing TyrRS-HA that cannot be alternatively spliced.…”

Section: Matrix Metalloproteinase Mmp2 Cleaves Tyrrs In Loser Cellsmentioning

confidence: 99%

Active JNK-dependent secretion of Drosophila Tyrosyl-tRNA synthetase by loser cells recruits haemocytes during cell competition

2015

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Cell competition is a process by which the slow dividing cells (losers) are recognized and eliminated from growing tissues. Loser cells are extruded from the epithelium and engulfed by the haemocytes, the Drosophila macrophages. However, how macrophages identify the dying loser cells is unclear. Here we show that apoptotic loser cells secrete Tyrosyl-tRNA synthetase (TyrRS), which is best known as a core component of the translational machinery. Secreted TyrRS is cleaved by matrix metalloproteinases generating MiniTyr and EMAP fragments. EMAP acts as a guiding cue for macrophage migration in the Drosophila larvae, as it attracts the haemocytes to the apoptotic loser cells. JNK signalling and Kish, a component of the secretory pathway, are autonomously required for the active secretion of TyrRS by the loser cells. Altogether, this mechanism guarantees effective removal of unfit cells from the growing tissue.

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Human tRNA synthetase catalytic nulls with diverse functions

Cited by 106 publications

References 34 publications

A Human Disease-causing Point Mutation in Mitochondrial Threonyl-tRNA Synthetase Induces Both Structural and Functional Defects

A Human Disease-causing Point Mutation in Mitochondrial Threonyl-tRNA Synthetase Induces Both Structural and Functional Defects

Identification and characterization of sORF-encoded polypeptides

Active JNK-dependent secretion of Drosophila Tyrosyl-tRNA synthetase by loser cells recruits haemocytes during cell competition

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