Understanding the Role of Cleavage in Cell-to-Cell Fusion of Mouse Hepatitis Virus

V, Sreeparna (2015) Understanding the Role of Cleavage in Cell-to-Cell Fusion of Mouse Hepatitis Virus. Masters thesis, Indian Institute of Science Education and Research Kolkata.

PDF (MS dissertation of Sreeparna V. (10MS76)) MS_thesis_Sreeparna_V_10MS76.pdf - Submitted Version Restricted to Repository staff only Download (1MB)

Abstract

The spike glycoprotein, expressed on the virion envelope, mediates many biological properties of Mouse hepatitis virus (MHV), including receptor attachment, fusion of viral and cell membranes during entry, cell-to cell fusion during viral spread, and immune activation. MHV-A59 and MHV2 are two closely related strains of MHV that differ in pathogenicity. Both MHV-A59 and MHV2 cause hepatitis and meningitis. In addition to this, MHV-A59 causes demyelination, axonal loss and syncytia formation in both in vitro and in vivo. The recombinant strains of MHV, RSA59 and RSMHV2 are isogenic (background is from MHV- A59 in both the strains) except for spike gene. The spike gene of RSA59 is from MHV-A59 and that of RSMHV2 is from MHV-2. RSA59 retains pathological properties of MHV- A59, including its ability to induce cell to cell fusion, whereas, RSMHV2 is unable to cause cell to cell fusion and impaired in causing demyeliantion. Previous studies showed that spike protein is one of the major determinants of MHV induced demyelination and also play a role in cell-to cell fusion but so far no studies are available to know whether spike protein mediated cell to cell fusion has any role in demyelination. RSA59 spike protein is cleaved into two subunits, S1 and S2 whereas RSMHV2 spike protein is not cleaved, making the cleavage domain a potential target that can account to the functional differences in fusogenicity. To study the role of cleavage domain of spike protein in cell-to cell fusion we are generating a recombinant RSA59 strain where the cleavage domain is exchanged with RSMHV2 by targeted RNA recombination. Our preliminary in vitro studies demonstrated that cleavage site mutated recombinant strain of RSA59 did not completely abolish its fusogenic properties. Experiments are in progress to characterize the newly generated strain and its pathogenecity in vivo in C57BL/6 mice.

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