Determinants of Mouse Hepatitis Virus Host Range Expansion

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ProQuest, 2006 - 113 pages
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The cross-species transmission of viruses is a poorly understood process, but never more relevant in light of the emergence of HIV and Severe Acute Respiratory Syndrome. Using the Coronavirus Mouse Hepatitis Virus (MHV), a persistent infection model and a mixed cell environment model (reflecting xenotransplantation conditions) were developed to study the molecular mechanisms of cross-species transmission. Mutant viruses isolated from both models displayed an expanded host range phenotype encompassing normally non-permissive cell lines from humans, monkeys, and hamsters. Mutations regulating host range were mapped to the Spike gene. The Spike protein is the principal mediator of virus-receptor binding and cellular entry. Several domains within the Spike protein have been associated with specific functions. They include: (1) a receptor binding domain mapped within the amino-terminal 330 amino acids, (2) an S1 cleavage subunit that is responsible for receptor interaction, and (3) an S2 cleavage subunit responsible for fusion and cellular entry.
  

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Contents

Chapter
1
Table Page 1 Representative Coronaviridae group members hosts and receptors
4
Figure Page 1 Comparison of MHV and SARSCoV genomes
6
Discontinuous transcription model
7
MHV Spike protein features
9
fMHV targeted RNA recombination
13
MHV host range model systems
17
Modification of the pMH 54 targeting vector
20
Immunofluorescence analysis of HepG2 cells
47
Receptor utilization by recombinant viruses
49
Amino acid substitutions in the Spike protein
68
CE AC AM 1 a receptor usage in STCEACAM1 a and DBT cells in the presence of blocking antibody CC1
69
Construction of C4 recombinant viruses
70
Construction of C4 S subdivision recombinants
71
Wild type and recombinant virus replication on human and hamster cell lines
72
HE recombinant virus selection and screening
73

Primers used in vector modification and recombinant construction
21
Recombinant virus construction strategy
23
Mutations in the Spike proteins derived from a persistent infection
42
Spike protein and substitution mapping
43
Mapping of host range determinants on human HepG2 cells
44
Fine mapping of host range determinants in S2 subunit
45
Recombinant virus replication in CHO and MCF7 cells
46
HE+ recombinant virus replication
74
Mapping of MHV host range determinants
75
CEACAM1 a receptor usage by recombinant viruses
76
Heparan sulfate as a mediator of infection
77
REFERENCES
87
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