trans-Golgi Retention of a Plasma Membrane Protein: Mutations in the Cytoplasmic Domain of the Asialoglycoprotein Receptor Subunit H1 Result in trans-Golgi Retention

• Published in: The Journal of cell biology 1995-07, Vol.130 (2), p.285-297
• Main Authors: Wahlberg, Johanna M., Geffen, Iris, Reymond, Françoise, Simmen, Thomas, Spiess, Martin
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Antibodies; Asialoglycoprotein Receptor; Asialoglycoproteins - metabolism; Base Sequence; Biochemistry; Cell Line; Cell lines; Cell membranes; Cells; Cellular biology; Complementary DNA; COS cells; Endoplasmic Reticulum - metabolism; Fluorescent Antibody Technique; Golgi Apparatus - metabolism; HeLa cells; Hepatocytes; Humans; Membrane proteins; Membrane Proteins - chemistry; Membrane Proteins - genetics; Membrane Proteins - metabolism; Molecular Sequence Data; Mutagenesis; Mutation; Proteins; Receptors; Receptors, Cell Surface - chemistry; Receptors, Cell Surface - genetics; Receptors, Cell Surface - metabolism; Transfection; Tumor Cells, Cultured
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.130.2.285

Unlike the wild-type asialoglycoprotein receptor subunit H1 which is transported to the cell surface, endocytosed and recycled, a mutant lacking residues 4-33 of the 40-amino acid cytoplasmic domain was found to be retained intracellularly upon expression in different cell lines. The mutant protein accumulated in the trans-Golgi, as judged from the acquisition of trans-Golgi-specific modifications of the protein and from the immunofluorescence staining pattern. It was localized to juxtanuclear, tubular structures that were also stained by antibodies against galactosyltransferase and γ-adaptin. The results of further mutagenesis in the cytoplasmic domain indicated that the size rather than the specific sequence of the cytoplasmic domain determines whether H1 is retained in the trans-Golgi or transported to the cell surface. Truncation to less than 17 residues resulted in retention, and extension of a truncated tail by an unrelated sequence restored surface transport. The transmembrane segment of H1 was not sufficient for retention of a reporter molecule and it could be replaced by an artificial apolar sequence without affecting Golgi localization. The cytoplasmic domain thus appears to inhibit interaction(s) of the exoplasmic portion of H1 with trans-Golgi component(s) for example by steric hindrance or by changing the positioning of the protein in the membrane. This mechanism may also be functional in other proteins.