The role of Npas4 in Neuroprotection and neurogenesis
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Date
2015
Authors
Choy, Fong Chan
Editors
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Lewis, Martin David
Koblar, Simon Andrea
Koblar, Simon Andrea
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Abstract
Neuronal Per-Arnt-Sim domain protein 4 (Npas4) is an activity-dependent transcription
factor belonging to the basic Helix-Loop-Helix-PAS family of regulatory proteins. Npas4 has
been demonstrated to play a role in regulating both inhibitory and excitatory synapse
formation in a neuronal cell type-specific manner and controlling the expression of brain-derived
neurotrophic factor, a neurotrophin that is important for neuronal survival,
differentiation and synaptic plasticity. Studies have also revealed a functional role for Npas4
in hippocampus- and amygdala-dependent learning and memory formation, as well as
cognitive and social neurobehaviour. Due to its central role in homeostasis of neuronal
excitation and inhibition, Npas4 has been linked to a host of psychiatric conditions such as
bipolar disorder, autism spectrum disorder and cognition-related disorders.
Upregulated Npas4 expression has been documented following various brain insults,
including ischaemia. Although previous studies have demonstrated a role for Npas4 in the
protection of neurons against several types of neurodegenerative insult, little is known
about its neuroprotective role in response to ischaemic brain injury. Chapter 2 demonstrates
that Npas4 has a neuroprotective role in ischaemia since Npas4 deficiency increased the
susceptibility of cultured neurons to cell death by oxygen and glucose deprivation and
aggravated the severity of brain injury after photochemical stroke in mice. Furthermore,
ablation of Npas4 caused an increase in activated astrocytes and microglia, pro-inflammatory
cytokines interleukin-6 and tumour necrosis factor alpha levels and a switch
from apoptotic to necrotic cell death following focal cerebral ischaemia. These findings
suggest that the mechanism underlying the in vivo protective effect of Npas4 after ischaemic
insult could be due to its capacity to limit progressive neurodegeneration and
neuroinflammation.
Npas4 is a brain-specific transcription factor whose expression has been reported to be
enriched in neurogenic regions of the brain, implicating a role for Npas4 in neurogenesis.
Using two independent in vitro models of neurogenesis, we recently demonstrated that
Npas4 expression is dynamic and highly regulated during neural differentiation of embryonic
stem cells (ESCs). However, the factors responsible for regulating Npas4 expression during
this process remain to be elucidated. Given that increasing evidence suggests that
microRNAs (miRNAs) play central roles in both embryonic and adult neurogenesis, we
reasoned that miRNAs are good candidates for the regulation of Npas4 expression during
neural differentiation of ESCs. Chapter 3 provides insight into the underlying mechanisms by
which Npas4 expression is regulated during neural differentiation of mouse ESCs (mESCs). In
this study, we utilized the 'next-generation' or 'deep' sequencing method to profile miRNA
expression during neural differentiation of mESCs. Two differentially expressed miRNAs were
identified to be able to significantly reduce reporter gene activity by targeting the Npas4
3'UTR, namely miR-744 and miR-224. More importantly, ectopic expression of these miRNAs
during neural differentiation resulted in downregulation of endogenous Npas4 expression.
Subsequent functional analysis revealed that overexpression of either miR-744 or miR-224
delayed neural differentiation, reduced GABAergic neuron production and inhibited neurite
outgrowth. Taken together, the data presented indicate that Npas4 not only plays a role at the early stages of neural differentiation of mESCs but may also, at least in part, influence
neuronal subtype specification and neurite development.
Given the significant role of miRNA in regulating gene expression, knowledge of the miRNA
profile across different time points during neural differentiation of mESCs will be crucial to
better understanding of the mechanisms governing neurogenesis. Chapter 4 further
examines the next-generation sequencing data generated from the second results chapter in
an attempt to identify isomiRs and novel miRNAs expressed during neural differentiation of
mESCs and assess their expression pattern throughout the differentiation process.
School/Discipline
School of Biological Sciences
Dissertation Note
Thesis (Ph.D.) -- University of Adelaide, School of Biological Sciences, 2015
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Copyright material removed from digital thesis. See print copy in University of Adelaide Library for full text.
This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at: http://www.adelaide.edu.au/legals
This electronic version is made publicly available by the University of Adelaide in accordance with its open access policy for student theses. Copyright in this thesis remains with the author. This thesis may incorporate third party material which has been used by the author pursuant to Fair Dealing exceptions. If you are the owner of any included third party copyright material you wish to be removed from this electronic version, please complete the take down form located at: http://www.adelaide.edu.au/legals