Differential recognition of tyrosine-based basolateral signals by AP-1B subunit μ1B in polarized epithelial cells

To investigate the importance of tyrosine recognition by the AP-1B clathrin adaptor subunit μ1B for basolateral sorting of integral membrane proteins in polarized epithelial cells, we have produced and characterized a mutant form of μ1B. The mutant (M-μ1B) contains alanine substitutions of each of the four conserved residues, which in the AP-2 adaptor subunit μ2 are critical for interacting with tyrosine-based endocytosis signals. We show M-μ1B is defective for tyrosine binding in vitro, but is nevertheless incorporated into AP-1 complexes in transfected cells. Using LLC-PK1 cells expressing either wild type or M-μ1B, we find that there is inefficient basolateral expression of membrane proteins whose basolateral targeting signals share critical tyrosines with signals for endocytosis. In contrast, membrane proteins whose basolateral targeting signals are distinct from their endocytosis signals (transferrin and low-density lipoprotein receptors) accumulate at the basolateral domain normally, although in a manner that is strictly dependent on μ1B or M-μ1B expression. Our results suggest that μ1B interacts with different classes of basolateral targeting signals in distinct ways.