Abstract

The long term goal of this project is to understand the molecular basis of the biological properties of the murine coronavirus, mouse hepatitis virus strain A59 (MHV-A59). MHV-A59 causes persistent demyelinating disease in the central nervous system of mice and readily establishes chronic, productive, non-lytic infections in glial cells in culture. Thus, the virus presents a model system in which to study viral persistence, tissue tropism and virus- induced demyelination. MHV infection of mice has been cited as a model for Multiple Sclerosis. We have been engaged in a detailed analysis of the molecular biology of MHV-A59. During the next five year period studies on the structure and function of the MHV-A59 non-structural genes, in particular the viral RNA-dependent RNA polymerase and the mechanism of transcription of mRNAs will be carried out. Antisera raised against bacterial/viral fusion proteins derived from cloned cDNA fragments from non-structural genes A,B,D, and E of MHV-A59 will be used to identify the proteins encoded by these genes. A map of the polypeptides encoded by the 24 kb putative polymerase gene A as well as the complete nucleotide sequence of the gene will be generated. As a long term goal the antisera will be used in an in vitro system to define the individual activities of the polymerase polypeptides. Studies will be carried out to test the leader-priming model for the initiation of viral mRNA transcription. Transfection of RNAs transcribed from pGEM vectors containing viral intergenic sequences upstream of the CAT gene will be carried out to determine whether these viral sequences can function as promoters for transcription by a leader primed mechanism. Site specific mutagenesis of leader and intergenic regions will identify nucleotide critical for transcription. In addition, RNA binding proteins will be identified by gel retardation assays and by assaying specific retention of leader intergenic RNA on protein blots. A long term goal will be to assemble a complete cDNA copy of the genome to be expressed as RNA to be used for mutagenesis to map the pathogenic properties of the virus.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI017418-12
Application #
3127225
Study Section
Virology Study Section (VR)
Project Start
1989-05-01
Project End
1994-04-30
Budget Start
1992-05-01
Budget End
1993-04-30
Support Year
12
Fiscal Year
1992
Total Cost
Indirect Cost

  Institution

Name
University of Pennsylvania
Department
Type
Schools of Medicine
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104

  Publications

Navas-Martin, Sonia; Brom, Maarten; Chua, Ming-Ming et al. (2007) Replicase genes of murine coronavirus strains A59 and JHM are interchangeable: differences in pathogenesis map to the 3'one-third of the genome. J Virol 81:1022-6
Navas-Martin, Sonia; Brom, Maarten; Weiss, Susan R (2006) Role of the replicase gene of murine coronavirus JHM strain in hepatitis. Adv Exp Med Biol 581:415-20
De Albuquerque, Nadine; Baig, Ehtesham; Ma, Xuezhong et al. (2006) Murine hepatitis virus strain 1 produces a clinically relevant model of severe acute respiratory syndrome in A/J mice. J Virol 80:10382-94
Zuo, Xun; Mattern, Michael R; Tan, Robin et al. (2005) Expression and purification of SARS coronavirus proteins using SUMO-fusions. Protein Expr Purif 42:100-10
Sperry, Steven M; Kazi, Lubna; Graham, Rachel L et al. (2005) Single-amino-acid substitutions in open reading frame (ORF) 1b-nsp14 and ORF 2a proteins of the coronavirus mouse hepatitis virus are attenuating in mice. J Virol 79:3391-400
Kazi, Lubna; Lissenberg, Arjen; Watson, Richard et al. (2005) Expression of hemagglutinin esterase protein from recombinant mouse hepatitis virus enhances neurovirulence. J Virol 79:15064-73
Weiss, Susan R; Navas-Martin, Sonia (2005) Coronavirus pathogenesis and the emerging pathogen severe acute respiratory syndrome coronavirus. Microbiol Mol Biol Rev 69:635-64
Navas-Martin, Sonia; Hingley, Susan T; Weiss, Susan R (2005) Murine coronavirus evolution in vivo: functional compensation of a detrimental amino acid substitution in the receptor binding domain of the spike glycoprotein. J Virol 79:7629-40
Chua, Ming Ming; MacNamara, Katherine C; San Mateo, Lani et al. (2004) Effects of an epitope-specific CD8+ T-cell response on murine coronavirus central nervous system disease: protection from virus replication and antigen spread and selection of epitope escape mutants. J Virol 78:1150-9
Navas-Martin, Sonia R; Weiss, Susan (2004) Coronavirus replication and pathogenesis: Implications for the recent outbreak of severe acute respiratory syndrome (SARS), and the challenge for vaccine development. J Neurovirol 10:75-85

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