Fig. 6. CNBD is essential for trafficking of ERG3. (A) Immunoblot analyses of cells transfected with ERG3_wt before and after treatment with PNGase, EndoH or tunicamycin. The mature (M) and immature (I) immunoreactive species are shown by arrows. Tunicamycin treatment was performed overnight 1 day after transfection. (B) Pulse-chase analysis of ERG3_wt at indicated chase times. Cells producing ERG3wt were labeled with radioactive cysteine and methionine, and chased at the indicated times. At the end of each chase interval, equal amounts of cell extracts were subjected to immunoprecipitation with an anti-HA antibody. The immunoprecipitated material was subjected to SDS-PAGE separation and autoradiography. Notice the gradual appearance of the mature band, which coincides with the disappearance of the immature species. (C) Immunoblot of cell extracts expressing ERG3 mutant variants. Cell lysates were subjected to EndoH (+) treatment and probed with an anti-HA antibody.