A Golgi retention signal in a membrane-spanning domain of coronavirus E1 protein

The G protein of vesicular stomatitis virus (VSV) is transported rapidly and efficiently to the plasma membrane in transfected cells, and much is known about its folding and oligomerization (7) . The single membrane-spanning domain of the G protein was replaced with that of ml from IBV El (Fig . 1 A) . The domain replacement was performed precisely using oligonucleotide-directed mutagenesis. The chimeric G protein, called Gml, was expressed transiently in Swift and Machamer Golgi Retention Signal COS cells using a SV-40-based vector (28) , or in HeLa cells, using a vaccinia virus T7 RNA polymerase expression system (10) . Gml was recognized by polyclonal antibodies to both the G ectodomain andthe cytoplasmic tail, and by two mAbs which recognize conformation-sensitive epitopes (Fig . 1 B, lanes 5-8) . In addition, Gml spanned the membrane since the cytoplasmic tail was susceptible to trypsin digestion in microsomal membranes (Fig . 1 B, lane 12) . These results indicated that ml functioned as a proper membrane-spanning domain in Gml, and that the chimeric protein was not grossly misfolded .