Identification of host and viral factors of arthritic alphavirus pathogenesis: the role of mannose binding lectin and the viral N-linked glycans

Arthritogenic alphaviruses such as Ross River virus (RRV) and chikungunya virus are mosquito-borne viruses that cause epidemics of debilitating myositis and polyarthritis in humans in various areas around the world. Studies conducted in a mouse model of RRV-induced disease have demonstrated a critical role for the inflammatory response in the development of disease. In particular, the host complement system contributes significantly to damage within target tissues through activation of CR3-bearing inflammatory cells. However, the precise mechanism and ligands leading to complement activation and disease following RRV infection are not known. In these studies, we have identified critical roles for the host innate immune protein mannose binding lectin (MBL) and the viral N-linked glycans in mediating complement activation and disease following RRV infection. Using mice deficient in MBL, we demonstrated that the MBL activation pathway of the host complement system is the primary pathway required for complement activation and disease following infection. MBL recognizes and binds to terminal carbohydrates, such as mannose found on glycosylated viral proteins or on infected cells. The RRV E2 envelope glycoprotein contains two N-linked glycosylation sites that are glycosylated with a combination of high mannose and complex glycans when replicating in mammalian cells. We hypothesized that MBL recognizes the E2 N-linked glycans to activate the complement system, leading to disease. Using a panel of RRV mutants lacking one or more envelope glycans, we have found that the RRV E2 N-linked glycans contribute to MBL binding to RRV infected cells and development of disease. Viruses lacking either E2 N-linked glycosylation sites cause reduced disease in mice, while a virus lacking both sites causes very mild disease. In addition, the role of the E2 glycans is independent of replication within host tissues and recruitment of inflammatory cells. Rather, the E2 glycans were required for MBL deposition and complement activation within target tissues in vivo. These results suggest that interactions between the viral N-linked glycans and the MBL pathway play a central role in development of severe alphavirus-induced arthritis and may be an effective target for therapeutic treatment in patients infected with RRV.Doctor of Philosoph