Use this URL to cite or link to this record in EThOS: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.805946
Title: Genetic characterisation of the Drosophila Mitochondrial Calcium Uniporter in physiological and neurodegenerative contexts
Author: Gleeson, Tom
Awarding Body: University of Cambridge
Current Institution: University of Cambridge
Date of Award: 2019
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Abstract:
Neurodegenerative conditions such as Alzheimer’s disease (AD) and Parkinson’s disease (PD) are a growing medical and social burden for which no disease-modifying therapies exist, necessitating greater understanding of their underlying pathobiology. Mitochondrial health and calcium signalling have come to the fore in the study of both conditions, and mitochondrial calcium dynamics, which play a crucial role in basal metabolism and cell death, have potential to be a key player in the disease process. Though mitochondrial calcium influx had long been observed through electrophysiology and other work, the genetic basis for this process has only recently been described. The uniporter is comprised of MCU, the pore-forming component; EMRE, which coordinates the complex architecture and is required for in vivo calcium uptake; and MICU1-3, which regulate the flow of ions. Animal models have emerged for some uniporter genes, but variable effects have left their physiological role uncertain. The genetic tractability of the Drosophila melanogaster model system makes it well situated to address this, and its short lifespan facilitates investigation of diseases of aging, but it has been little utilised in the study of the uniporter. Here, I complete a full suite of genetic tools for interrogating mitochondrial calcium uniporter components conserved in Drosophila melanogaster, through molecular cloning, P-element transposition, and CRISPR-mediated targeted mutagenesis. Knockouts of MCU and EMRE, both essential for in vivo uniporter activity, are largely tolerated in the fly, though lifespan is reduced, especially for MCU. Metabolic features of these mutants also diverge. The loss of the regulatory MICU1 results in developmental lethality, preceded by organismal dysfunction. Crucially, this was not rescued by MCU or EMRE knockout, indicating the presence of a uniporter-independent role of MICU1. Knockouts of MICU3 were viable but displayed defects associated with tissues in which the gene is more expressed. In addition to characterisation of the physiological role of these genes, I crossed them to Drosophila models of neurodegenerative disease, primarily Pink1 loss. Reduction of MCU strikingly rescued Pink1-associated deficits, with more variable rescue against parkin, demonstrating some specificity for mitophagy-independent Pink1 functions. MCU loss also markedly improved a mutant amyloid expressing model of Alzheimer’s disease. As well as shedding light on the physiological requirements of mitochondrial calcium uniporter components, my work therefore argues for a subset of neurodegenerative conditions being amenable to modification of mitochondrial calcium influx. Further work should build on this potential target to attempt to ameliorate these otherwise intractable diseases.
Supervisor: Whitworth, Alexander J. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.805946  DOI:
Keywords: mitochondria ; Drosophila ; MCU ; EMRE ; micu1 ; micu3 ; Alzheimer's disease ; Parkinson's disease ; calcium ; genetics
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