Characterization of the Nipah Virus P gene Products And the Innate Immune Responses of Nipah Virus Infected Endothelial and Neuronal Cells 公开

Lo, Michael K. (2009)

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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


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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