trans-Golgi retention of a plasma membrane protein: mutations in the cytoplasmic domain of the asialoglycoprotein receptor subunit H1 result in trans-Golgi retention

Here we describe mutants of a cell surface receptor, the ASGP receptor subunit HI, that pass the quality control mechanisms of the ER, but are retained in a later compartment of the secretory pathway and do not reach the plasma membrane. The site of accumulation was identified as the trans-Golgi/TGN based on biochemical and morphological evidence for the mutant Hl(A4-33A). The retained protein was modified by the trans-Golgi-specific enzymes sialyltransferase and (when tagged with an appropriate acceptor sequence) tyrosylprotein sulfotransferase. By indirect immunofluorescence microscopy, the mutant was found in juxtanuclear structures that are distinct from endosomes and lysosomes, and that are also stained with antibodies directed against galactosyltransferase and ~,-adaptin. Furthermore, the effects of nocodazole, okadaic acid, brefeldin A, and chloroquine on the distribution of H1 (A4-33A) in MDCK cells were consistent with a trans-Golgi localization. were fixed without (A and C) or with permeabilization (B and D) and stained with antibodies specific for subunit H1 (A and B) or for subunit H2 (C and D). Bar, 10 Ixm.