QUALITY-CONTROL IN THE SECRETORY PATHWAY - THE ROLE CALRETICULIN, CALNEXIN AND BIP IN THE RETENTION OF GLYCOPROTEINS WITH C-TERMINAL TRUNCATIONS

Unlike properly folded and assembled proteins, most misfolded and inco mpletely assembled proteins are retained in the endoplasmic reticulum of mammalian cells and degraded without transport to the Golgi complex . To analyze the mechanisms underlying this unique sorting process and its fidelity, the fate of C-terminally truncated fragments of influen za hemagglutinin was determined An assortment of different fragments w as generated by adding puromycin at low concentrations to influenza vi rus-infected tissue culture cells. Of the fragments generated, < 2% wa s secreted, indicating that the system for detecting defects in newly synthesized proteins is quite stringent. The majority of secreted spec ies corresponded to folding domains within the viral spike glycoprotei n. The retained fragments acquired a partially folded structure with i ntrachain disulfide bonds and conformation-dependent antigenic epitope s. They associated with two lectin-like endoplasmic reticulum chaperon es (calnexin and calreticulin) but not BiP/GRP78. Inhibition of the as sociation with calnexin and calreticulin by the addition of castanospe rmine significantly increased fragment secretion. However, it also cau sed association with BiP/GRP78. These results indicated that the assoc iation with calnexin and calreticulin was involved in retaining the fr agments. They also suggested that Bip/GRP78 could serve as a backup fo r calnexin and calreticulin in retaining the fragments. In summary, th e results showed that the quality control system in the secretory path way was efficient and sensitive to folding defects, and that it involv ed multiple interactions with endoplasmic reticulum chaperones.