Misregulation of stress granules by tRNA synthetases as a potential pathogenic mechanism in Charcot-Marie-Tooth neuropathy

Dominant mutations in regions encoding aminoacyl-tRNA synthetase enzymes (aaRSs) are associated with hereditary Charcot-Marie-Tooth (CMT) peripheral neuropathy, a neurodegenerative disease characterized by degradation of the peripheral neurons. While aaRSs functions primarily in the aminoacylation of tRNAs with their cognate amino acids, recent evidence indicates that many of these pathogenic variants are not associated with significant loss of aminoacylation activity. Recent mRNA binding assays in yeast models have demonstrated a previously unknown mRNA binding activity of several aaRSs. Like many mRNA binding proteins, during stress aaRSs are found in large assemblies of protein and RNA called stress granules. The existence of this secondary function of tRNA synthetases informs a new direction in the investigation of the pathogenic mechanism for these variants. Plasmids containing mutations in the yeast histidyl-tRNA synthetase gene (HARS) associated with the development of CMT were obtained from the Antonellis Lab at the University of Michigan Medical School. These plasmids were transformed into yeast strains in which the genomic copy of histidyl-tRNA synthetase had been deleted. The resulting strains were transformed with plasmids encoding a fluorescently labeled stress granule marker proteins for microscopic analysis. Stress granule formation was analyzed in these HARS mutants, as abnormalities in granule formation are commonly associated with the misregulation of translation and the development of neurodegenerative diseases. This work will inform the hypothesis that misregulation of translation or stress granule formation by HARS could lead to the development of CMT in individuals with these deleterious mutations