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

To generate the eDNA of H1(A4-33A) Ts, a variant of Hl(A4-33A) tagged with a tyrosine-sulfation peptide at the carboxy terminus, the 3' portion (BamHl-EcoRI) of the eDNA of Hl(A4-33A) was replaced by that of H1TM described by Leitinger et al. (1994) . To construct Hl(A4-33A) myc, a variant containing a carboxy-terminal c-myc epitope tag, the tyrosine-sulfation tag sequence (KpnI-SacI) of the H1 rs eDNA was replaced by two complementary oligonucleotides, CGAACAAAAGTTGA-TITCTGAAGAAGACTrGAACTGAGCT and CAGTTCAAGTCT-TCITCAGAAATCAACITI'rGTTCGGTAC, which encode the peptide sequence EQKLISEEDLN followed by a stop codon and produce terminal protruding ends corresponding to a KpnI and a SacI site at the 5' and 3' end, respectively. All mutagenized segments were confirmed by DNA sequencing.