The dileucine motif within the tail of MPR46 is required for sorting of the receptor in endosomes

The cytoplasmic tail of MPR46 carries a leucine-based motif that is require d for the sorting of lysosomal enzymes by the receptor. In addition, it is one of three independent, but functionally redundant, internalization signa ls present in the cytoplasmic tail of MPR46, We have analyzed a mutant of M PR46, in which the dileucine pair was replaced by alanines (MPR46 LL/AA) wi th respect to its intracellular distribution and trafficking. Ultrastructur al analysis of cells expressing the MPR46 LL/AA mutant revealed that the su bstitution of the dileucine pair causes a shift of the receptor distributio n from the TGN, where it is packaged into AP1-containing vesicles, to vesic ular structures distributed throughout the cytoplasm. The vesicles could be identified as early endosomes with internalized BSA-gold and rab5 as marke rs. By analyzing the receptor trafficking biochemically, we found that retu rn of the LL/AA mutant receptor from the plasma membrane/endosome pool back to the TGN was impaired, while recycling from endosomes to the plasma memb rane was enhanced. In conclusion, our data indicate that the dileucine moti f in the MPR46 tail is required for a sorting event in endosomes.