A humanized yeast model reveals dominant-negative properties of neuropathy-associated alanyl-tRNA synthetase mutations

Abstract

Aminoacyl-tRNA synthetases (ARSs) are ubiquitously expressed, essential enzymes that ligate tRNA molecules to their cognate amino acids. Heterozygosity for missense variants or small in-frame deletions in five ARS genes causes axonal peripheral neuropathy, a disorder characterized by impaired neuronal function in the distal extremities. These variants reduce enzyme activity without significantly decreasing protein levels and reside in genes encoding homo-dimeric enzymes. These observations raise the possibility of a dominant-negative effect, in which non-functional mutant ARS subunits dimerize with wild-type ARS subunits and reduce overall ARS activity below 50%, breaching a threshold required for peripheral nerve axons. To test for these dominant-negative properties, we developed a humanized yeast assay to co-express pathogenic human alanyl-tRNA synthetase (AARS1) mutations with wild-type human AARS1. We show that multiple loss-of-function, pathogenic AARS1 variants repress yeast growth in the presence of wild-type human AARS1. This growth defect is rescued when these variants are placed in cis with a mutation that reduces dimerization with the wild-type subunit, demonstrating that the interaction between mutant AARS1 and wild-type AARS1 is responsible for the repressed growth. This demonstrates that neuropathy-associated AARS1 variants exert a dominant-negative effect, which supports a common, loss-of-function mechanism for ARS-mediated dominant peripheral neuropathy.