Ligand-Bound GeneSwitch Causes Developmental Aberrations inDrosophilathat Are Alleviated by the Alternative Oxidase

Abstract

Culture of Drosophila expressing the steroid-dependent GeneSwitch transcriptional activator under the control of the ubiquitous α-tubulin promoter was found to produce extensive pupal lethality, as well as a range of dysmorphic adult phenotypes, in the presence of high concentrations of the inducing drug RU486. Prominent among these was cleft thorax, seen previously in flies bearing mutant alleles of the nuclear receptor Ultraspiracle and many other mutants, as well as notched wings, leg malformations, and bristle abnormalities. Neither the α-tubulin-GeneSwitch driver nor the inducing drug on their own produced any of these effects. A second GeneSwitch driver, under the control of the daughterless promoter, which gave much lower and more tissue-restricted transgene expression, exhibited only mild bristle abnormalities in the presence of high levels of RU486. Coexpression of the alternative oxidase (AOX) from Ciona intestinalis produced a substantial shift in the developmental outcome toward a wild-type phenotype, which was dependent on the AOX expression level. Neither an enzymatically inactivated variant of AOX, nor GFP, or the alternative NADH dehydrogenase Ndi1 from yeast gave any such rescue. Users of the GeneSwitch system should be aware of the potential confounding effects of its application in developmental studies.

Developmental arrest in Drosophila melanogaster caused by mitochondrial DNA replication defects cannot be rescued by the alternative oxidase

“…Heat production by mitochondrial uncoupling is to date only detectable in AOX-expressing cells in the presence of antimycin A 32 , but the lowering of ROS apparently occurs even under normal physiological conditions when OXPHOS is properly functioning, at least in adult flies 33 . In addition, the fact that AOX can rescue developmental defects not related, a priori , to mitochondrial dysfunction, such as cleft thorax, notched wings, leg malformations, and bristle abnormalities in Drosophila 20 , indicates that yet unknown cellular mechanism(s) is(are) operating when AOX is present in the mitochondria of higher animals.…”

“…AOX function has also been reported to provide some benefit in other models of mitochondrial disorders, such as fly lines with knockdown of pol γ 17 or with mutations in dj-1β , the homologue of the human Parkinson’s disease gene DJ1 12 , and in sesB , the homologue of another PEO gene, ANT1 18 . In addition, the rescue by AOX of Drosophila models of Alzheimer’s disease and mid-line closure defects suggests that RC dysfunction and/or excess ROS production are not only involved, but also appear in early stages of these pathogeneses 19 , 20 .…”

Alternative NAD(P)H dehydrogenase and alternative oxidase: Proposed physiological roles in animals

“…For instance, AOX from Ciona intestinalis showed rescuing of various pathological symptoms in fruit flies with modelled Alzheimer disease (El-Khoury et al, 2016). As mentioned above, AOX was surprisingly able to rescue developmental abnormalities related to thoracic closure in fruit fly (Andjelković et al, 2016). It is also planned to apply AOX towards ischemia injuries (Rustin and Jacobs, 2009;Mills et al, 2016).…”

“…Thus during regeneration, NDH2 and AOX would enable oxidation of reducing equivalents formed in Krebs cycle in maturating mitochondria. Reports on the involvement of AOX in development, particularly in apoptosis and cellular differentiation, were recently published (Andjelković et al, 2016). It was found that ectopic expression of AOX from Ciona intestinalis may rescue developmental abnormalities in fruit flies by probable modulation of the c-Jun N-terminal kinase (JNK) signalling pathway (Andjelković et al, 2016).…”

“…Reports on the involvement of AOX in development, particularly in apoptosis and cellular differentiation, were recently published (Andjelković et al, 2016). It was found that ectopic expression of AOX from Ciona intestinalis may rescue developmental abnormalities in fruit flies by probable modulation of the c-Jun N-terminal kinase (JNK) signalling pathway (Andjelković et al, 2016). Thus, the consequences of the presence of AOX in the RS may spread beyond the reaction it catalyses.…”