The role of the ataxia protein sacsin in mitochondrial function

Supervisor: Dr Paul Chapple
Internal co-supervisor: Dr Tristan McKay
External co-supervisor: Professor Michael Duchen (University College London)

Project to be 50% funded by the Ataxia Charlevoix-Saguenay Foundation (Montreal)

Mitochondria constantly fuse and divide, mixing mitochondrial membranes and contents. These dynamic processes are essential for cellular health, with disruption of mitochondrial plasticity occurring in metabolic and neuronal diseases.

Mutations in the protein sacsin cause the inherited neurodegenerative disease Autosomal Recessive Spastic Ataxia of Charlevoix-Saguenay (ARSACS). We have shown sacsin contains functional domains linking it to proteostasis (Parfitt et al 2009, Human Molecular Genetics). We have also localised sacsin to mitochondria in neurons and demonstrated that its knockdown alters mitochondrial morphology. Specifically, cells with reduced levels of sacsin have elongated and more interconnected mitochondria. Moreover, sacsin interacts with the mitochondrial fission protein Dpr1. Together these data support a role for sacsin in mitochondrial fission. A manuscript arising from this work has been accepted by PNAS subject to revision.

Through this studentship we will investigate the consequences of loss of sacsin function for mitochondrial network in human neurons. We will also further test the hypothesis that disruption of mitochondrial dynamics underlies ARSACS. From collaboration with clinical geneticist at the University of Nijmegen (Netherlands) we have obtained ARSACS patient fibroblasts and demonstrated they do not express sacsin. Further ARSACS patient fibroblasts will be available from Dr Paola Giunti, a neurologist at the UCL Institute of Neurology. We will compare mitochondrial networks and function in these patient fibroblasts to control. Moreover, we will generate iPS cells from patient and control fibroblasts and differentiate them into neuronal cell types (Dr Mckay is expert in these techniques). These human model systems will be used to analyse the consequences, for mitochondria, of loss of sacsin function. The overall aim will be pursued through four key objectives that will determine:

1. The impact of sacsin knockdown on mitochondrial bioenergetic function.
2. The role of sacsin in mitochondrial fusion, fission and mitophagy.
3. The impact of sacsin knockdown on mitochondrial trafficking.
4. The impact of reduced sacsin expression on mitochondrial calcium handling and free radical generation.

The predominant experimental approach used will be fluorescence imaging by laser scanning confocal microscopy. Professor Duchen is expert in the application of fluorescence imaging to mitochondrial biology.

This studentship will help Dr Chapple cement international collaborations with researchers in Canada and Holland. It will also utilize a number of contemporary cell biology techniques and develop new expertise in the WHRI. For example, expanding our repertoire of approaches to monitor mitochondrial function through interaction with Professor Duchen (this will be of use to other researchers in Endocrinology such as Dr Metherell and Dr Storr). The cutting edge nature of the project will make it highly attractive to high quality students.

In summary this research will define how sacsin modulates mitochondrial dynamics and function, and potentially the cellular mechanism by which mutations in sacsin cause ARSACS. It will also further elucidate our understanding of mitochondrial quality control in neurons.