Mutation of the Conserved Polyadenosine RNA binding protein, ZC3H14/dNab2, Impairs Neural Function in Drosophila and Humans Open Access

Abstract

Every cell contains the same DNA sequence. Yet tight
regulation of gene expression occurs such that different tissues of a multicellular
organism achieve gene expression profiles in cell-type specific,
spatial and temporal manners. While this regulation of gene expression can occur at the
level of transcription and epigenetic modification, post-transcriptional regulatory
mechanisms are also critical. Between transcription and translation, many processes are
dedicated to ensure proper processing and maturation of transcripts and thus allow for
proper regulation of translation. RNA-binding proteins play
critical roles in achieving this regulation as highlighted by
many human diseases. Interestingly, mutations in genes encoding RNA-binding proteins
that are ubiquitously expressed and play important roles for overall RNA metabolism in
all cell types result in tissue-specific phenotypes. Why certain tissues are more sensitive
to defects in general RNA-binding protein functions is unclear. Studies addressing both
developmental and tissue-specific functional characterization of critical RNA-binding
proteins will allow for better understanding of human disease biology.
We show for the first time that mutations in the human ZC3H14 gene lead to non-
syndromic autosomal recessive intellectual disability (NS-ARID) and use Drosophila
melanogaster to model key aspects of the disease. Here we describe the first
identification and characterization of dNab2, a Drosophila orthologue of ZC3H14/Nab2
class of Cys3 His (CCCH) tandem zinc finger (ZnF) polyadenosine RNA-binding protein.
dNab2 is essential for development and required in neurons for normal locomotion and
flight. Biochemical and genetic data indicate that dNab2 restricts bulk poly(A) tail length
in vivo, suggesting that this function may underlie its role in development and disease.
Furthermore, we define the role of dNab2 in controlling locomotor activity and memory
formation in the Drosophila mushroom bodies (MBs), a highly specialized structure
involved in higher cognitive functions and locomotion. Finally, using a genetic modifier
screen, we identify putative dNab2 targets and/or interacting proteins that modulate
dNab2-mediated neuronal function. These studies reveal a conserved requirement for
ZC3H14/dNab2 in the metazoan nervous system and allow for future studies on the
molecular mechanisms underlying ZC3H14-associated human intellectual disability.

Table of Contents

Table of Contents
Chapter 1 Introduction
1

Introduction
2
I. Post-transcriptional regulation of gene expression in eukaryotes
3
II. Control of poly(A) tail length as a key step in post-transcriptional regulation of gene
expression
6
III. RNA-binding proteins implicated in human disease
14
IV. Drosophila melanogaster as a model system
19
V. Scope and significance of the dissertation
24
Figures
28
References
36
Chapter 2 Mutation of the Conserved Polyadenosine RNA binding protein,
ZC3H14/dNab2 , Impairs Neural Function in Drosophila and Humans
51
Introduction
54
Results
56
Discussion
63
Figures
65
Experimental procedures
82
References
87
Chapter 3 Defining the role of dNab2 in the Drosophila mushroom bodies
93
Introduction
94
Results
97
Discussion
104
Figures
107
Experimental procedures
117
References
119
Chapter 4 A genetic-modifier screen identifies factors that functionally interact with
the Drosophila dNab2 gene
123
Introduction
124
Results
126
Discussion
133
Figures
136
Experimental procedures
148
References
150
Chapter 5 Conclusion and Discussion
153
Conclusion
154
A model
154
Remaining questions
157
Future directions
159

About this Dissertation

Rights statement

• Permission granted by the author to include this thesis or dissertation in this repository. All rights reserved by the author. Please contact the author for information regarding the reproduction and use of this thesis or dissertation.

School	Laney Graduate School
Department	Biological and Biomedical Sciences
Subfield / Discipline	Genetics and Molecular Biology
Degree	PhD
Submission	Dissertation
Language	English
Research Field	Biology, Genetics Biology, Cell
Keyword	Drosophila development RNA-binding proteins polyadenylation intellectual disability
Committee Chair / Thesis Advisor	Moberg, Kenneth H, Emory University Corbett, Anita, Emory University
Committee Members	Lucchesi, John, Emory University Kelly, William G, Emory University Boss, Jeremy, Emory University Bassell, Gary, Emory University

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