Targeting of protein ERGIC-53 to the ER/ERGIC/cis-Golgi recycling pathway

We have shown that virtually the entire cytoplasmic domain is required for targeting of ERGIC-53, but is the cytoplasmic domain of ERGIC-53 also sufficient for correct targeting of a reporter protein within the ER-ERGIC-cis-Golgi cycle? By double immunofluorescence microscopy the pattern of L4T4C53 was compared with endogenous ERGIC-53 16 h after transfection. L4T4C53 showed mostly an ER pattern with a strong nuclear ring clearly distinct from endogenous ERGIC-53 (Fig. 8 B, a and b) , while localization of a construct containing both the cytoplasmic and the lumenal domain of ERGIC-53, L53T4C53, was indistinguishable from GM (not shown). This difference in localization may be due to the reduced influence of the RSQQE determinant in the CD4-chimera L4T4C53. Indeed a CD4-chimera lacking the KKFF-retrieval signal, L4T4C53 (K-4stop), acquired 32% endo H resistance after 1 h of chase (not shown) considerably more than the 12% for the corresponding mutation in L53T4C53 (Fig. 6 , line 5). We conclude therefore that the presence of the ERGIC-53 lumenal domain is required for a functional RSQQE determinant.