Deletions into an NH2-terminal hydrophobic domain result in secretion of rotavirus VP7, a resident endoplasmic reticulum membrane glycoprotein

A series of plasmids containing mutations in the VP7 gene has also been constructed (see Fig. 2 ). The deletions were constructed in order to study the role of the hydrophobic domains in VP7 synthesis. The location of VP7 expressed from these plasmids in COS 7 cells was also examined by immunofluorescence. The deletion mutant 1-14 displayed extensive ER and perinuclear staining in regions distinct from that seen for R-WGA (Fig. 5, c and d) , similar to the wildtype. In this mutant, the first ATG codon was deleted so that only the second initiation codon and the hydrophobic region following it could be used (Fig. 2) . Another mutant studied, 2-8, in which the sequence following the first ATG codon was modified (see Materials and Methods), showed results analogous to the wild-type (Fig. 5 , e and f). In contrast, VP7 distribution in the mutants 42-61, 43-61, and 47-61, in which distal parts of the second hydrophobic domain were removed (Fig. 2) , showed reticular staining and a distinct perinuclear distribution which coincided precisely with, or was a subset of, the R-WGA staining pattern (Fig. 5, g-l) .