Functional and genetic analysis of alpha-synuclein and huntingtin in Drosophila melanogaster

211 Pages·2016·5.06 MB·English

Functional and genetic analysis of alpha-synuclein
and huntingtin in Drosophila melanogaster
Gonçalo Manuel Poças
Dissertation presented to obtain the Ph.D degree in Biochemistry
Instituto de Tecnologia Química e Biológica António Xavier | Universidade Nova de Lisboa
Oeiras,
December, 2015
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Functional and genetic analysis of
alpha-synuclein and huntingtin in
Drosophila melanogaster
Gonçalo Manuel Poças
Dissertation presented to obtain the Ph.D degree in Biochemistry
Instituto de Tecnologia Química e Biológica António Xavier | Universidade Nova de Lisboa
Oeiras, December, 2015
Title: Functional and genetic analysis of alpha-synuclein and huntingtin in
Drosophila melanogaster

Dissertation presented to obtain the Ph.D. degree in Biochemistry

Author: Gonçalo Manuel Poças

Cell Signaling in Drosophila Laboratory
Instituto de Tecnologia Química e Biológica
Universidade Nova de Lisboa
Av. da República
Estação Agronómica Nacional
2780-157 Oeiras
Portugal

Front cover picture: image obtained by confocal microscopy of a Drosophila
brain expressing alpha-synuclein (green) in the dopaminergic neurons marked
by a specific antibody for Tyrosine Hydroxylase (blue).

Copyright © 2015 by Gonçalo Manuel Poças
All Rights Reserved
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I declare that the work presented in this thesis, except where otherwise stated,
is based on my own research and it was supervised by Doctor Pedro Manuel
Domingos. The work was mainly performed in Instituto de
Tecnologia Química e Biológica, Universidade Nova de Lisboa.

I am grateful for the financial support provided by Fundação para a Ciência e
Tecnologia (grant reference SFRH/BD/61477/2009).

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Aos meus pais

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Table of contents

Acknowledgements ......................................................................................... xi
Summary ...................................................................................................... xiii
Sumário .......................................................................................................... xv
Thesis publications .................................................................................... xvii
List of acronyms ........................................................................................ xviii

Chapter I ..................................................................................................................... 1
General Introduction

1.1. Human neurodegenerative diseases ....................................................................... 3
1.2. Parkinson’s disease ................................................................................................ 8
1.2.1. Epidemiology, symptoms and general molecular features .......................... 8
1.2.2. α-Synuclein: a major player in Parkinson’s disease .................................. 14
1.2.3. The vulnerability of dopaminergic neurons in Parkinson’s disease .......... 17
1.3. Huntington’s disease ............................................................................................ 21
1.3.1. Epidemiology, symptoms and general molecular features ........................ 21
1.3.2. Huntingtin: the monogenic cause of Huntington’s disease ........................ 22
1.3.3. Medium spiny neurons vulnerability in Huntington’s disease .................. 25
1.4. Modeling Parkinson’s and Huntington’s diseases in Drosophila ........................ 27
1.5. Main goals ........................................................................................................... 35
1.6. References ........................................................................................................... 36

Chapter II ................................................................................................................ 49
The subcellular localization and axonal transport of α-synuclein in a Drosophila
model for Parkinson’s disease

2.1. Abstract ............................................................................................................... 51
2.2. Introduction ......................................................................................................... 52
2.3. Results
2.3.1. The wild-type form of α-syn accumulates in the synaptic terminals of
Drosophila photoreceptors ................................................................................... 56
2.3.2. The A30P mutant version of α-syn is mislocalized in the Drosophila
photoreceptors ..................................................................................................... 57
2.3.3. No differences were detected in the aggregation state of the wild-type and
the A30P mutant version of α-syn ...................................................................... 59
2.3.4. Identification of α-syn WT and A30P protein interactors by
co-immunoprecipitation and mass spectrometry analysis .................................. 62
2.3.5. Tomosyn, Spaghetti Squash, and Synaptotagmin 4 are modulators α-syn
axonal transport and subcellular localization....................................................... 66
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