Project Outline

Mitochondrial DNA (mtDNA) is a small genome with a big impact on health. Over 350 pathogenic mtDNA mutations have been linked to a spectrum of mitochondrial diseases that affect 1 in 5,000 individuals. Intriguingly, the type and severity of symptoms often vary immensely among individuals carrying the same mtDNA mutation. As mitochondrial function requires the coordinated expression of nuclear and mitochondrial genes, many unexplained variations in the penetrance of mtDNA-linked disorders could be caused by mito-nuclear allelic interplays (1, 2). Yet, how nuclear polymorphisms influence mitochondrial performance in a given mtDNA background remains largely unknown.

Genetic tools recently developed in Drosophila allow us to isolate mtDNA mutants in animals (3). Using these mutants, we found cases where mito-nuclear epistasis modulates the severity of mtDNA-linked disorders. In particular, we performed a population screen and showed that a temperature-sensitive mtDNA mutant with compromised complex IV activity (mt:CoIts) could be robustly rescued by nuclear polymorphisms presented in 2 out of 192 strains. The rescued flies show significantly improved energy production, lifespan and mobility. We sequenced and localised the responsible polymorphisms to a small region on the 2nd chromosome. Based on the nature of sequence variations, we shortlisted seven genes as potential suppressors. This project aims to identify and characterise how nuclear suppressor polymorphisms modulate the pathogenic expression of mtDNA mutations. There are two objectives: 1) map nuclear polymorphisms that alleviate mt:CoIts defects by genetic crosses and mutant isolation; and 2) investigate the nature of rescue by cellular and biochemical approaches. If time allows, the student will also use a similar approach to probe tissue-specific mito-nuclear epistasis using mtDNA mutants with testis-only defects (e.g. male sterile).

The outputs of this project will increase our understanding of genome evolution and provide valuable information for the management of mitochondrial diseases. In particular, identifying and understanding how nuclear-encoded proteins lessen defects induced by mtDNA mutations can lead to new targets and strategies for therapeutic intervention. A better understanding of the impact of mito-nuclear interactions is also invaluable for predicting prognosis and tissue-specific presentation in patients with mitochondrial disorders. Furthermore, with the advent of a mitochondrial replacement therapy that pairs nuclear DNA with different mitochondrial genomes to create “three-parent babies”, the fitness consequences of mito-nuclear interactions are of immediate medical relevance.

References

1. Xu H, DeLuca SZ, O'Farrell PH. Manipulating the metazoan mitochondrial genome with targeted restriction enzymes. Science 321:575-7, 2008.
2. Wolff JN, Ladoukakis ED, Enríquez JA, Dowling DK. Mitonuclear interactions: evolutionary consequences over multiple biological scales. Philos Trans R Soc Lond B Biol Sci. 369:20130443, 2014.
3. Carelli V, Giordano C, d'Amati G. Pathogenic expression of homoplasmic mtDNA mutations needs a complex nuclear-mitochondrial interaction. Trends Genet. 19:257-62, 2003.

Techniques

• Drosophila pushing/genetic crosses, DNA/RNAseq, Genetic manipulation.
• Biochemical assays including Blue Native PAGE, Western Blot and Co-Immunoprecipitation, Seahorse & High-resolution respirometry for cellular metabolism analysis.
• Mass spectrometric analysis.
• Confocal imaging & EM imaging.