The signal for clathrin-mediated endocytosis of the paramyxovirus SV5HN protein resides at the transmembrane domain-ectodomain boundary region

The hemagglutinin-neuraminidase (HN) glycoprotein of the paramyxovirus SV5 is internalized from the cell surface via clathrin-coated pits. However, th e cytoplasmic domain of SV5 HN does not contain a previously characterized internalization motif. A cell-surface-expressed chimeric protein (APK), con sisting of the cytoplasmic tail, transmembrane (TM) domain, and 12 residues of the ectodomain of HN joined to the cytoplasmic protein pyruvate kinase is internalized, indicating that the N-terminal region of HN contains an in ternalization signal. Although SV5 HN is internalized at a rate similar to that of influenza virus hemagglutinin (HA) mutant Y543, which contains a de generate tyrosine-based signal in its cytoplasmic tail, the elimination of the majority of the HN cytoplasmic tail, or substitution of the HN TM domai n with leucine residues, did not affect the rate of HN internalization. The HN protein of the closely related virus, Newcastle disease virus (NDV), is not internalized from the cell surface. Working under the usual convention that the TM domain consists of the hydrophobic residues bounded by two cha rged residues, analysis of internalization of mutant and chimeric NDV HN mo lecules indicates that the first seven SV5 HN ectodomain residues are criti cal for internalization of HN. A glutamic acid residue (537) that abuts thi s presumptive HN TM domain/ectodomain boundary is important for SV5 HN inte rnalization. (C) 1999 Academic Press.