The glycobiology of the Nipah virus fusion and attachment proteins

Nipah virus (NiV) causes severe and often fatal febrile encephalitis. The fusion (F) and attachment (G) envelope glycoprotein of NiV mediate endothelial syncytia formation, a prominent hallmark of NiV infections. There is no efficacious treatment for Nipah virus infection, thus we set out to establish targets for treatment. Given that attachment of the virus to the cell and fusion of the viral membrane with the cell membrane are the first steps in viral infection, we focused our efforts on the study of Nipah virus F and G proteins.;NiV-F and NiV-G were codon optimized and we established that these versions of the genes produced more protein and led to higher syncytia formation than their wild-type counterparts. A NIV-G/Fc fusion protein was created, which bound to permissive but not to non-permissive cells and inhibited NiV fusion. Moreover the optimized genes were used in mutational analysis of all potential N-linked glycan sites on the glycoproteins and established that the glycosylation state of these proteins was important. Furthermore, NiV envelope-mediated cell-cell fusion is blocked by galectin-1, an endogenous lectin. Inhibition is specific to the Paramyxoviridae family and requires the physiologic dimeric form of galectin-1. Galectin-1 also aberrantly oligomerizes NiV-F and NiV-G.;In addition, we determined that ephrinB2 is the cellular receptor for the Nipah virus. EphrinB2 binds to NiV-G and ephrinB2-Fc effectively blocks NiV fusion and entry; moreover, transfection of ephrinB2 into non-permissive cells renders them permissive for NiV fusion and entry. EphrinB2 is expressed on endothelial cells and neurons consistent with the known cellular tropism for NiV.;Lastly, one mechanism of galectin-1 inhibition of NiV fusion was identified. Galectin-1 did not block NiV-G binding to the cell surface or to recombinant ephrinB2. However, galectin-1 specifically prevented the endocytosis of NiV-F. In addition, pulse chase and cell biological tracking experiments demonstrated that galectin-1 decreased the amount of cleaved NiV-F and specifically disrupted the colocalization of NiV-F with an early endosomal marker.;Thus we have developed a system for studying the attachment and fusion proteins of the NiV, which has led to the identification of the cellular receptor and an inhibitor of fusion. All of this will help to shed light on the pathobiology of NiV infection, and spur the rational development of effective therapeutics.