Coronaviruses are a common cause of respiratory infection in man and cause many diseases in domestic animals, including respiratory and enteric infections, neurological disease and hepatitis. Because of the difficulty of isolating coronaviruses from clinical specimens, the full spectrum of coronavirus-induced disease in man is not yet known. We will explore interactions of a murine coronavirus with different host cells to determine the molecular basis for its strong species and tissue tropisms. We will analyze the structural and functional domains of the 3 major proteins in mouse hepatitis virus, using a large number of monoclonal antibodies which we have developed as well as antibodies to synthetic peptides mimicking important regions of viral peptides. We will determine the roles in virus replication of virus-specific RNA-binding proteins and a newly discovered 140K virion-associated protein. We will explore the assembly of these enveloped virions at intracytoplasmic membranes by biochemical, immunological and electron microscopic techniques. We will make use of virus mutants defective in assembly, infection of hybridoma cells producing antibodies to viral proteins, and microinjection of monoclonal antibodies into living cells. Using a variety of susceptible and resistant cell types, we will analyze the cellular mechanisms for restriction of coronavirus replication. We will compare virus attachment and penetration in these different cell types, and use a new in vitro assay to learn whether differences in transcription of viral RNA and translation of viral proteins can account for differential susceptibility of cells to coronavirus infection. We will determine the role in coronavirus maturation of host cell lipids and host-dependent proteolytic cleavage of a viral glycoprotein.
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