Notably, when expressed in Drosophila, YARS1 E196K- similarly to the aminoacylation-compromised YARS1 G41R and YARS1 153-156delVKQV- induces phenotypes recapitulating hallmark features of the human pathology, including motor deficits, electrophysiological neuronal dysfunction and axon terminal degeneration 15. All five known CMT-causing mutations are in the catalytic domain of the protein 2, 3, and yet YARS1 E196K does not impair the enzyme kinetics 12. In this study, we focused on YARS1, a cytoplasmic tRNA ligase with additional nuclear roles in transcriptional regulation and response to oxidative stress 13, as well as extracellular signaling 14. An overarching and potentially common mechanistic hypothesis explaining the neuropathology shared by these enzymes is currently lacking and this is hampering the development of effective therapeutics. The structural relaxation of mutant GARS1 and AARS1 leads to a neomorphic interaction with the transmembrane receptor Neuropilin 1 8, 11, while the conformational opening caused by three different CMT mutations in YARS1 increases the binding affinity to TRIM28 12, compared with the wild type proteins. Notably, the CMT mutations in GARS1 8, YARS1 9, HARS1 10, and AARS1 11, cause 3D-conformational opening that exposes sequences buried within the wild-type enzymes, thereby impacting their interaction properties. To search for commonalities in the mode of action of the mutant synthetases, systematic structural studies have been performed. CMT mutations could disrupt those non-aminoacylation activities or generate completely novel neurotoxic properties of the affected synthetases. Rather, a growing list of diverse noncanonical functions is being described for these enzymes 2, 6, 7. Extensive evidence from us and others demonstrates that, at least for YARS1, glycyl-tRNA synthetase (GARS1) and alanyl-tRNA synthetase (AARS1), the CMT phenotype cannot be ascribed to a simple loss of aminoacylation (reviewed in refs. How monoallelic genetic defects in such ubiquitous enzymes can cause selective damage of the peripheral nerves is not fully understood. The pathology is mostly restricted to the axons of the peripheral nerves that degenerate in a length-dependent, dying-back fashion upon disease progression. Dominant mutations in tyrosyl-tRNA synthetase (YARS1) and six other synthetases cause different subtypes of Charcot-Marie-Tooth disease (CMT) 2– 4, the most common and currently incurable inherited peripheral neuropathy 5.

These ubiquitous enzymes are responsible for the continuous and correct charging of tRNAs with their cognate amino acids during protein biosynthesis (canonical function) 1. Protein translation is a fundamental cellular process in which aminoacyl-tRNA synthetases play a major role. Hence, in this work, we show that YARS1 is an evolutionary-conserved F-actin organizer which links the actin cytoskeleton to tRNA-synthetase-induced neurodegeneration. Similar beneficial effects are observed in flies expressing a neuropathy-causing glycyl-tRNA synthetase. Genetic modulation of F-actin organization improves hallmark electrophysiological and morphological features in neurons of flies expressing CMT-causing YARS1 mutations. Biochemical studies uncover yet unknown actin-bundling property of YARS1 to be enhanced by a CMT mutation, leading to actin disorganization in the Drosophila nervous system, human SH-SY5Y neuroblastoma cells, and patient-derived fibroblasts. By an unbiased genetic screen in Drosophila, we link YARS1 dysfunction to actin cytoskeleton organization. Loss of aminoacylation is not required for their pathogenicity, suggesting a gain-of-function disease mechanism. Relevant code and classifiers (e.g., ImageJ macro scipts) are available from the corresponding author on reasonable request.ĭominant mutations in tyrosyl-tRNA synthetase (YARS1) and six other tRNA ligases cause Charcot-Marie-Tooth peripheral neuropathy (CMT). Source data are provided with this paper. Databases and softwares relevant to the study include ImageJ FlyBase PANTHER the quantitative proteomics software package MaxQuant ClustalOmega DroID. The human biomaterials are available subject to MTA. All other relevant data are available from the corresponding author on reasonable request. The mass spectrometry data are deposited at the PRIDE database under accession number PXD037630. 1, 2, 4, 5, 7– 10 are provided as a Source Data file. GUID: 1D93E3F5-3EBA-4A8A-AC0A-AFB1EC2C2062 Data Availability StatementĪll data generated and/or analyzed during this study are included in this article (and its supplementary files).

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