(A) The sequences of the calsequestrin 2 (CASQ2) 81-nt sequence from the region of overlap between ORF-1 and ORF-2 subjected to mutation. The multiple aspartate residues of ORF-1 are shown in bold and italics and the potential initiation methionine residues of ORF-2 are shown in bold and underlined. The wild-type unaltered sequence is shown (CASQ2 wt) together with the CASQ2 D/E mutant overlap in which the multiple aspartate residues were altered to glutamate residues and the CASQ2 D/E plus mutant in which nine aspartate residues at the amino terminal of the overlap region were maintained followed by the glutamic acid motif used in the CASQ2 D/E construct. (B) Detection of GFP and CAT protein in lysates of cells transfected with expression plasmids by Western blot. GFP-specific and CAT-specific monoclonal antibodies were used to detect protein expression. Cells were transfected with the plasmid construct indicated. Lanes on the left of each pair are the constructs identified and lanes on the right of each pair are the appropriate STOP mutant. Controls were GFP and CAT protein standards and lysate from cells transfected with empty expression vector, as indicated.
