LOCALIZATION OF BUNYAMWERA BUNYAVIRUS G1-GLYCOPROTEIN TO THE GOLGI REQUIRES ASSOCIATION WITH G2 BUT NOT WITH NSM

The Bunyamwera bunyavirus (BUN) M RNA genome segment encodes three pro teins, two glycoproteins termed G1 and G2 and a non-structural protein called NSm, in the form of a polyprotein precursor that is cotranslat ionally cleaved to give the mature proteins. Indirect immunofluorescen ce experiments have shown that these proteins localize to the Golgi co mplex in BUN-infected cells. We have used a recombinant vaccinia virus (vTF7-3), which expresses bacteriophage T7 RNA polymerase, to drive t he expression of plasmids containing either the entire BUN M segment c DNA or fragments that encode the G1, G2 and NSm proteins separately un der control of the T7 promoter. After transfection of these plasmids i nto vTF7-3-infected cells, correctly sized and processed proteins were detected by immunoprecipitation with BUN-specific antibodies. Immunof luorescence experiments showed that G1, G2 and NSm localized to the Go lgi when transiently expressed from the full-length cDNA. When G2 or N Sm were expressed separately they also localized to the Golgi, but whe n G1 was expressed alone a staining pattern typical for the endoplasmi c reticulum was obtained. However coexpression of G2 and G1 from indep endent plasmids resulted in G1 localizing to the Golgi. In contrast tr anslocation of G1 to the Golgi was not observed when G1 was coexpresse d with NSm, although NSm itself was still detected in the Golgi. Simil ar results were obtained when the proteins were expressed from transfe cted plasmids containing the G2-, NSm- or G1-coding sequences under co ntrol of the cytomegalovirus immediate-early promoter. The localizatio n of G1 to the Golgi when coexpressed with G2 was confirmed by the los s of endoglycosidase H (endo H) sensitivity of G1 after approximately 60 min in a pulse-chase experiment; G1 remained sensitive to endo H wh en expressed either alone or in combination with NSm. These results su ggest that G2 contains the Golgi targeting and/or retention signals an d that G1 has to interact with this protein to localize to this cellul ar compartment.