Chemical cross-linking of proteins of Semliki Forest virus: virus particles and plasma membranes from BHK-21 cells treated with colchicine or dibucaine.

Abstract

Chemical cross-linking of the proteins of Semliki Forest virus has been performed in virus particles and in baby hamster kidney-21 (BHK-21) cells infected with Semliki Forest virus. Most of the studies were done with the reversible cross-linkers dimethyl 3,3'-thiobis(propionimidate) and dithiobis(succinimidyl propionate). The identity of the cross-linked species was determined by two-dimensional electrophoresis. The results with virus particles showed extensive cross-linking of the nucleocapsid proteins and the formation of dimers of the two large envelope glycoproteins (E1 and E2). Similar patterns for the cross-linked virus proteins were observed in plasma membranes isolated from BHK-21 cells infected with Semliki Forest virus. No cross-linking of the third envelope glycoprotein (E3) was observed. Also, there was no evidence for significant cross-linking between host and virus proteins. The addition of colchicine, a drug that disrupts microtubules, to infected BHK-21 cells had no effect on the cross-linking of virus proteins in the plasma membrane. In contrast, dibucaine, a local anesthetic, greatly inhibited the formation of envelope dimers (E1-E2) in plasma membranes, but not in virus particles. The implication of these results for the involvement of the cytoskeletal system in the morphogenesis of Semliki Forest virus is discussed.