technical knockout, a Drosophila Model of Mitochondrial Deafness

Abstract

Mutations in mtDNA-encoded components of the mitochondrial translational apparatus are associated with diverse pathological states in humans, notably sensorineural deafness. To develop animal models of such disorders, we have manipulated the nuclear gene for mitochondrial ribosomal protein S12 in Drosophila (technical knockout, tko). The prototypic mutant tko25t exhibits developmental delay, bang sensitivity, impaired male courtship, and defective response to sound. On the basis of a transgenic reversion test, these phenotypes are attributable to a single substitution (L85H) at a conserved residue of the tko protein. The mutant is hypersensitive to doxycyclin, an antibiotic that selectively inhibits mitochondrial protein synthesis, and mutant larvae have greatly diminished activities of mitochondrial redox enzymes and decreased levels of mitochondrial small-subunit rRNA. A second mutation in the tko gene, Q116K, which is predicted to impair the accuracy of mitochondrial translation, results in the completely different phenotype of recessive female sterility, based on three independent transgenic insertions. We infer that the tko25t mutant provides a model of mitochondrial hearing impairment resulting from a quantitative deficiency of mitochondrial translational capacity.

Mitochondrial Protein Translation: Emerging Roles and Clinical Significance in Disease

“…Analysis of the patient showed a homozygous C-to-T substitution at nucleotide 331 of the MRPS16 cDNA (Miller et al, 2004). Similar results showed that in Drosophila, a missense mutation in mitochondrial ribosomal protein S12 prevented ribosomal proteins from assembling into active ribosomes, resulting in a significant reduction in 12S rRNA transcripts (Toivonen et al, 2001). Another mitochondrial ribosomal SSU protein mutation occurs in MRPS22.…”

<b>Adenylate kinase 2 deficiency limits survival and </b><b>regulates various genes during larval stages of </b><b><i>Drosophila melanogaster </i></b>

“…Furthermore, mutations in mitochondrial rRNA transfer and mitochondrial aminoacyl tRNA synthetase genes were identified as causative of deafness (30,31). Mutation of the mitochondrial ribosomal protein S12 (technical knockout, tko) was also reported as a cause of deafness in fruit flies (32). Taken together, these observations strongly indicate functional associations between defects of mitochondrial translation machinery and hearing loss.…”

Nuclear genetic background influences the phenotype of theDrosophila tko25tmitochondrial protein-synthesis mutant

“…Courtship was analyzed essentially as previously (Toivonen et al 2001), in batches of 6 pairs of adult male and virgin female 3 day-old flies of a given genotype. For experiments at different temperatures, virgin flies were kept at the experiment temperature until the experiment was started.…”