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REGULATION OF PROTEIN AGGREGATION BY ARFAPTIN2 IN AMYOTROPHIC LATERAL SCLEROSIS

Mohammedeid, Aida (2015) REGULATION OF PROTEIN AGGREGATION BY ARFAPTIN2 IN AMYOTROPHIC LATERAL SCLEROSIS. PhD thesis, University of Sheffield.

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Abstract

Amyotrophic lateral sclerosis (ALS) is a devastating, adult onset motor neuron disease (MND) that has no effective treatment to date. The current study investigates the possibility of targeting protein aggregation pathway for treatment. Modulation of this pathway is approached through targeting Arfaptin2 protein. The C-terminal of Arfaptin2 (HC-ARFIP2), a dominant negative form of Arfaptin2, has been shown to have a neuroprotective property that maintains the proteasome activity and induces degradation of misfolded proteins. We thus proposed that the HC-ARFIP2 improves neuronal survival in ALS via maintaining the proteasomal pathway, which in turn will decrease neuron toxicity and improve neuronal survival. Expression of HC-ARFIP2 in primary motor neuron cultures using LV-based vector, improved motor neuron survival significantly. The prosurvival effect was observable even in cells treated with H2O2 in both SOD1G93A transgenic and non-transgenic cells. A further investigation on the pathway of which HC-ARFIP2 exerts it neuroprotective effect showed that HC-ARFIP2 induces Akt phosphorylation by decreasing its dominant negative modulator, PTEN. In addition, protein degradation pathway-markers (p62, LC3II, ULK1) showed significant changes in response to HC-ARFIP2 expression. Furthermore, Arfaptin2 showed colocalisation with SOD1 and overexpression of FL-ARFIP2 caused aggregates formation in HEK293T cells compared to HC-ARFIP2 expression that maintained the cytoplasmic distribution of SOD1. A scAAV9-based vector was produced to replicate these observations in vivo. The desired viral titre could not be obtained within the time course of the PhD. In conclusion, the study presented here has provided a proof of concept that Arfaptin2 is involved in protein aggregation in ALS. In addition, HC-ARFIP2 expression can improve motor neuron survival in vitro through activation of Akt and proteasome activity.

Item Type: Thesis (PhD)
Academic Units: The University of Sheffield > Faculty of Medicine, Dentistry and Health (Sheffield)
The University of Sheffield > Faculty of Medicine, Dentistry and Health (Sheffield) > Medicine (Sheffield)
Identification Number/EthosID: uk.bl.ethos.657011
Depositing User: Dr Aida Mohammedeid
Date Deposited: 14 Jul 2015 11:15
Last Modified: 03 Oct 2016 13:15
URI: http://etheses.whiterose.ac.uk/id/eprint/9377

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