Characterization of the Nipah Virus P gene Products And the Innate Immune Responses of Nipah Virus Infected Endothelial and Neuronal Cells Public
Lo, Michael K. (2009)
Abstract
Characterization of the Nipah Virus P gene Products
And the Innate Immune Responses of
Nipah Virus Infected Endothelial and Neuronal
Cells
By Michael K. Lo
Nipah virus (NiV) is a highly pathogenic paramyxovirus which
frequently causes
fatal encephalitis in humans, and the
molecular mechanisms of NiV pathogenesis are unclear. Endothelial
cells and neurons
are important cellular targets in the pathogenesis of this disease.
The goals of this
dissertation are to characterize the expression of NiV
phosphoprotein (P)-derived gene
products in the context of infection, and to characterize the
endothelial and neuronal cell
innate immune responses against NiV infection.
In this study, our sequence analysis of multiple cloned mRNAs
from infected
cells showed that henipavirus P gene mRNA editing frequencies are
higher than
those reported for most other paramyxoviruses. Antisera generated
against synthetic
peptides from the P, V, W, and C proteins of NiV were able to
detect all four proteins in
NiV infected cells and in purified virions. In infected Vero cells,
the W protein was
detected in the nucleus while P, V, and C were found in the
cytoplasm. The W protein
co-immunoprecipitated with karyopherin alpha3.
Although plasmid expression studies indicated the ability of
individual NiV P gene products to antagonize the innate antiviral
response in several
cell lines, it is not known whether live NiV infection of
physiologically relevant cellular
targets reflect those results. Little has been done
in regards to the molecular mechanisms of vasculitis seen in human
cases. In this study,
we characterized the growth kinetics and the innate immune
responses of primary
endothelial cells and a neuronal cell line. NiV infected
endothelial cells produced a
functional IFN-β response, which correlated with a
differential localization of
the NiV W protein when compared with infected neuronal cells, which
lacked any
detectable antiviral response. We demonstrated that NiV infection
of endothelial
cells induced a significant increase of inflammatory chemokines
secreted into the cellular
supernatant, which induced a corresponding increase in
monocyte
and T-lymphocyte chemotaxis. This study is the first in
vitro characterization of the
innate immune response against NiV infection in physiologically
relevant cell types.
Table of Contents
Table of Contents
Page
Chapter 1. Literature Review 1
Introduction 2
Epidemiology 3
Virus Reservoir 5
Clinical presentation & pathological manifestations 6
Classification and Morphology 10
The Genome Organization of NiV 12
General Overview of the NiV Replication Cycle 13
NiV Membrane Proteins 16
The Ribonucleoprotein Complex 24
The Innate Cellular Antiviral Response 31
Virus-Host interactions of the NiV P gene products 33
References 37
Chapter 2. Determination of Henipavirus P gene mRNA Editing55
Frequencies and Detection of the C, V, and W Proteins
of Nipah Virus in Virus-Infected Cells
Abstract 56
Introduction 57
Results 60
Discussion 65
Materials and Methods 70
References 79
Chapter 3. Characterization of the growth kinetics and the93
innate immune response against Nipah virus
in endothelial cells and neurons
Abstract 94
Introduction 95
Results 98
Discussion 103
Materials and Methods 108
References 115
Chapter 4. Discussion and Conclusions 128
References 139
About this Dissertation
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