Wednesday, 7 December 2016

Selective removal of deletion-bearing mitochondrial DNA in heteroplasmic Drosophila

http://www.nature.com/articles/ncomms13100

Nikolay P. Kandul, Ting Zhang, Bruce A. Hay & Ming Guo

Mitophagy is the process by which mitochondria are engulfed and degraded within the cell. It has long been thought that mitophagy has a role to play in quality control in the mitochondrial population, however it has remained unclear whether the effect can selectively degrade faulty organelles, or instead is an unbiased process, in vivo. A potential confounding effect is cell division where, simply by dividing, mutants can be driven to fixation through stochastic effects.

To address this question, the authors developed a Drosophila model which can inducibly generate a mitochondrial mutation which would otherwise be lethal if present at birth in the whole-body. This is done through the inducible expression of a mitochondrially-targetted restriction enzyme which cleaves mtDNA in Drosophila in two places, creating a 2584 bp deletion which disrupts or removes several important mitochondrial genes. The authors were able to induce expression of this restriction enzyme in a non-essential, energy-intense, post-mitotic tissue, namely the indirect flight muscle. This is an ideal tissue to study mitophagy, since its energy requirements imply the need for a healthy mitochondrial population, and is non-dividing so does not suffer from confounding effects from the stochastic nature of mtDNA dynamics.

Flies tended to accumulate ~76% heteroplasmy in the mitochondrial deletion by day 10 after hatching, stabilizing thereafter, with no large difference in mtDNA copy number. The flies had similar flight performances to wild-type animals, suggesting that the tissue may withstand high levels of heteroplasmy without phenotypic consequences.

The authors probed the effects of modulating the expression of genes which have been thought to play a role in mitophagy, and measured the resultant heteroplasmy. They investigated Atg1, Atg8a, Pink1 and Parkin, which all had the expected effects on heteroplasmy. Parkin overexpression caused ~71% reduction in heteroplasmy, and Atg1 caused ~72% reduction, these genes having the largest effect sizes. The authors found that inhibition of mitochondrial fusion through MFN silencing had a modest effect on heteroplasmy reduction (37%), which is expected if defective mitochondria are not allowed to re-enter the mitochondrial network. Interestingly, inhibiting ATP synthase from hydrolysing ATP and therefore maintaining mitochondrial membrane potential, through expression of ATPIF1, had a synergistic effect with MFN silencing, resulting in a 64% effect size.  This suggests that mitochondria with mutated mtDNAs may attempt to cheat the mitophagic system by consuming ATP to maintain their membrane potential and avoid detection.



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Thoughts:

These results show that mitophagy can be a selective process, and may be induced to have greater effect sizes. The key question is, if mitophagy is able to clear mitochondrial mutants, why do we see them at all in the wild-type case? What is the tradeoff that keeps mitophagy low? It would be interesting to see the lifespan of these flies upon induction of mitophagy. Are they more susceptible to other pathologies e.g. cancer or aging?

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