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

The mutant proteins were all inserted into the membrane and glycosylated as shown by immunoprecipitation from [ 3 IS]cysteine-labeled transfected COS cells (Fig. 5 B) . Mutation of Asn22 (N122) reduced the amount of fully glycosylated protein (most had one N-linked oligosaccharide instead of two), perhaps by conformationally altering the aminoterminal domain. Localization of the mutant proteins was determined by indirect immunofluorescence microscopy. With the exception of the Leu30 to Gln change (LQ3o), all of the mutations introduced into the full-length El protein appeared to hinder transport out of the ER (Fig. 6 ) . This is seen by the reticular staining pattern which includes nuclear envelope . This suggested that the Ile mutations might be disrupting proper folding ofthe El protein, perhaps by interfering with association of ml with the other two membranespanning domains, or with insertion into the membrane . In contrast, the LQ3o mutation had no apparent effect on targeting of El. Since mutating the polar residues to Ile prevented the exit of the mutant proteins from the ER, we were not able to assess their effects on retention in the Golgi complex .