Characterization of plasma membrane-associated proteins from Aedes albopictus mosquito (C6/36) cells that mediate West Nile virus binding and infection

Abstract

This study isolated and characterized the West Nile virus (WNV) putative receptor molecule(s) from Aedes albopictus mosquito (C6/36) cells. The binding of WNV to C6/36 cells was saturated with 5000 particles per cell. The entry of WNV into C6/36 cells was strongly inhibited when pretreated with proteinase K and to a lesser extent with sodium periodate. However, pretreatment of C6/36 cells with phospholipases, glycosidases, heparinases and neurimidase had no effect on virus entry. By using virus overlay protein blot assay, WNV was observed to bind to the 140-kDa, 95-kDa, 70-kDa and 55-kDa plasma membrane-associated molecules isolated from C6/36 cells. Murine antibodies generated against the 95-kDa and 70-kDa membrane proteins effectively blocked WNV, Japanese encephalitis virus (JEV) and Dengue virus (DV) serotype 2 infection in C6/36 cells. In addition, the binding of the recombinant-WNV envelope domain III protein to C6/36 cells can be inhibited by the anti-95-kDa and anti-70-kDa membrane protein antibodies. These data strongly supported the possibility that the 95-kDa and 70-kDa plasma membrane-associated proteins are part of a receptor complex for mosquito-borne flaviviruses (WNV, JEV and DV) on mosquito cells.