Identification of a precursor for one of the semliki forest virus membrane proteins

Abstract

We are studying the envelope of Semliki forest virus (SFV) as a model for membrane structure and biogenesis. Semliki forest virus and other group A arboviruses acquire their envelopes by budding through the host cell plasma membrane [ 1] . The viral envelope resembles the host cell plasma membrane in its lipid composition [2] , but has a much simpler protein composition: one protein with an apparent molecular weight of about 50,000 has been found in the membrane [3]. In Sindbis virus this protein has recently been split into two bands (Er and Ez) using discontinuous sodium dodecyl sulphate (SDS) polyacrylamide gel electrophoresis [4]. Both of these polypeptides contain carbohydrate. Studies on SFV protein formation are facilitated by the fact that virus infection effectively shuts off host cell protein synthesis [5]. Attempts to find out whether the structural polypeptides of the virus are translated as primary gene products or in precursor form(s) have so far been inconclusive [6-91. Our previous studies have suggested a precursor role for a non-structural glycopolypeptide (NSP68) which is found in extracts of infected cells [lo]. This polypeptide has an apparent molecular weight of 68,000 in 5% acrylamide SDS-gels. Here we show that SFV contains two envelope polypeptides, El and Ea , and that these have different primary structures. We also show that NSP68 is a precursor for the envelope polypeptide Ez, which is confirmed by the isolation of a temperaturesensitive mutant of SFV, in which precursor cleavage is blocked.