Multiple C-terminal motifs of the 46-kDa mannose 6-phosphate receptor tailcontribute to efficient binding of medium chains of AP-2 and AP-3

The interaction of adaptor protein (AP) complexes with signal structures in the cytoplasmic domains of membrane proteins is required for intracellular sorting. Tyrosine- or dileucine-based motifs have been reported to bind to medium chain subunits (mu) of AP-1, AP-2, or AP-3. In the present study, m e have examined the interaction of the entire 67-amino acid cytoplasmic dom ain of the 46-kDa mannose B-phosphate receptor (MPR46-CT) containing tyrosi ne- as well as dileucine-based motifs with mu2 and mu 3A chains using the y east two-hybrid system. Both mu2 and mu 3A bind specifically to the MPR46-C T. In contrast, mu 3A fails to bind to the cytoplasmic domain of the 300-kD a mannose 6-phosphate receptor. Mutational analysis of the MPR46-CT reveale d that the tyrosine-based motif and distal sequences rich in acidic amino a cid residues are sufficient for effective binding to mu2. However, the dile ucine motif was found to be one part of a consecutive complex C-terminal st ructure comprising tyrosine and dileucine motifs as well as clusters of aci dic residues necessary for efficient binding of mu 3k Alanine substitution of 2 or 4 acidic amino acid residues of this cluster reduces the binding to mu 3A much more than to mu2. The data suggest that the MPR46 is capable of interacting with different AP complexes using multiple partially overlappi ng sorting signals, which might depend on posttranslational modifications o r subcellular localization of the receptor.