| Nipah virus is a highly lethal emerging paramyxovirus that spreads systemically through the endothelial cells and targets the lung and brain tissue. Thus, infections cause respiratory complications and a severe febrile encephalitis. Progress is being made in development of antivirals and vaccines that may slow the pathogenesis of the disease. However, it is critical that we understand the molecular mechanisms of viral replication as they may hold the key to better therapeutics. In our studies, we sought to characterize the function of two structural proteins, M and P, involved in multiple steps of the viral life cycle.;We have defined the matrix protein, M, as the critical driving force for virion assembly and egress from the cell. When a sequence, YMYL, within NiV M was mutated, virus-like particle release was abolished and M was relocalized to the nucleus, but the mutant M proteins retained oligomerization activity. When YMYL was fused to a late domain mutant of the Ebola virus VP40 matrix protein, VP40 budding was restored. These results suggest that the YMYL sequence may act as a trafficking signal and a late domain for NiV M.;The phosphoprotein, P, is a cofactor for the viral polymerase and an antagonist of the interferon system. We have defined domains required for these functions and found that mutations can abolish interaction with the critical transcription factor STAT1 white leaving replication function intact. Thus, these mutations have been incorporated into a recombinant Nipah virus using our newly established reverse genetics system. The mutant viruses are unable to block the cellular response to interferon and, as such, should be sensitive to its antiviral effects. Additionally, investigation of the STAT1-binding domain (residues 114-140) determined that when fused to GST, this small domain prevented STAT1 activation by multiple stimuli. This direct interaction of 114-140 with STAT1 is sufficient to block phosphorylation of STAT1 in vitro.;In sum, this work focuses on functional domains within the M, P, V and W proteins required for Nipah virus assembly, viral RNA synthesis and inhibition of the host antiviral response. |
