title: Defective valyl-tRNA synthetase hampers the mitochondrial respiratory chain in Neurospora crassa.
abstract: Respiratory chain deficiency can result from alterations in mitochondrial and/or cytosolic protein synthesis due to the dual genetic origin of mitochondrial oxidative phosphorylation. In the present paper we report a point mutation (D750G) in the bifunctional VARS (valyl-tRNA synthetase) of the fungus Neurospora crassa, associated with a temperature-sensitive phenotype. Analysis of the mutant strain revealed decreased steady-state levels of VARS and a clear reduction in the rate of mitochondrial protein synthesis. We observed a robust induction of the mitochondrial alternative oxidase with a concomitant decrease in the canonical respiratory pathway, namely in cytochrome b and aa3 content. Furthermore, the mutant strain accumulates the peripheral arm of complex I and depicts decreased levels of complexes III and IV, consistent with severe impairment of the mitochondrial respiratory chain. The phenotypic alterations of the mutant strain are observed at the permissive growth temperature and exacerbated upon increase of the temperature. Surprisingly, glucose-6-phosphate dehydrogenase activities were similar in the wild-type and mutant strains, whereas mitochondrial activities for succinate dehydrogenase and alternative NADH dehydrogenases were increased in the mutant strain, suggesting that the VARSD-G mutation does not affect overall cytosolic protein synthesis. Expression of the wild-type vars gene rescues all of the mutant phenotypes, indicating that the VARSD-G mutation is a loss-of-function mutation that results in a combined respiratory chain deficiency.
authors.affiliation: IBMC (Instituto de Biologia Molecular e Celular), Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal. mduarte@ibmc.up.pt
authors.forename: Margarida
authors.initials: M
authors.lastname: Duarte
authors.forename: Arnaldo
authors.initials: A
authors.lastname: Videira
mesh_terms: D000595:Amino Acid Sequence; D004579:Electron Transport / Q000235:genetics; D008928:Mitochondria / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D009492:Neurospora crassa / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D017354:Point Mutation / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014637:Valine-tRNA Ligase / Q000172:deficiency* / Q000235:genetics / Q000502:physiology*
chemical_list: D014637:Valine-tRNA Ligase