King's College London

Mitochondrial retrograde signalling communicates the functional status of mitochondria to the cell by regulating nuclear gene transcription. Mitochondrial dysfunction is strongly associated with several neurodegenerative diseases, but little is known about mitochondrial retrograde signalling in the nervous system. Mitochondria and the endoplasmic reticulum (ER) share physical and functional links, but the impact of mitochondrial dysfunction on ER biology has been underexplored. In this thesis, I harnessed the array of genetic and functional tools available in the fruit fly, Drosophila melanogaster, to investigate mitochondrial retrograde signalling via the ER in the nervous system. I employed overexpression of mitochondrial transcription factor A (TFAM) as a model of mitochondrial dysfunction. The mitochondrial network was fragmented in the cell bodies of motor neurons overexpressing TFAM, and the morphology of the ER was also changed. There was an increase in the number of candidate ER-mitochondrial contact sites. Mitochondrial Ca2+ levels were substantially increased by TFAM overexpression, as were the levels of cytosolic Ca2+ in stimulated neurons. Mitochondrial dysfunction activated the ER stress signalling pathway, the unfolded protein response (UPR), in Drosophila motor neurons. Genetic manipulation of the UPR revealed that multiple components modify the function of Drosophila motor neurons with mitochondrial dysfunction. In particular, knock-down of the UPR transcription factors ATF4, XBP1 and ATF6 suppressed the neuronal functional impairments caused by mitochondrial dysfunction. Characterisation of the transcriptional response to mitochondrial dysfunction suggested that ATF4 is a key regulator of retrograde signalling in the Drosophila nervous system; 58% of genes that were significantly regulated by TFAM overexpression, were similarly regulated by overexpression of ATF4. Knock-down of ATF4 in conjunction with TFAM overexpression reversed the transcriptional changes caused by TFAM overexpression for 30% of genes. Therefore, I have established that the ER UPR is a mitochondrial retrograde signalling pathway in Drosophila neurons, and importantly, that targeting the UPR can alleviate the effects of mitochondrial dysfunction in neurons.