A PDZ-interacting domain in CFTR is an apical membrane polarization signal

Abstract

Polarization of the cystic fibrosis transmembrane conductance regulator (CFTR), a cAMP-activated chloride channel, to the apical plasma membrane of epithelial cells is critical for vectorial transport of chloride in a variety of epithelia, including the airway, pancreas, intestine, and kidney. However, the motifs that localize CFTR to the apical membrane are unknown. We report that the last 3 amino acids in the COOH-terminus of CFTR (T-R-L) comprise a PDZ-interacting domain that is required for the polarization of CFTR to the apical plasma membrane in human airway and kidney epithelial cells. In addition, the CFTR mutant, S1455X, which lacks the 26 COOH-terminal amino acids, including the PDZ-interacting domain, is mispolarized to the lateral membrane. We also demonstrate that CFTR binds to ezrin-radixin-moesin-binding phosphoprotein 50 (EBP50), an apical membrane PDZ domain-containing protein. We propose that COOH-terminal deletions of CFTR, which represent about 10% of CFTR mutations, result in defective vectorial chloride transport, partly by altering the polarized distribution of CFTR in epithelial cells. Moreover, our data demonstrate that PDZ-interacting domains and PDZ domain-containing proteins play a key role in the apical polarization of ion channels in epithelial cells.

Ezrin–Radixin–Moesin-Binding Phosphoprotein 50 Is Expressed At the Apical Membrane of Rat Liver Epithelia

“…In other expression studies, EBP50 has been implicated in directly modifying CFTR Cl Ϫ channel activity 45 and directing the cellular distribution of CFTR. 46 Consequently, it will be important to further characterize the effects of PDZ1 expression on the location and function of CFTR as well as other cAMP-regulated anion channels.…”

Myosin VI Regulates Endocytosis of the Cystic Fibrosis Transmembrane Conductance Regulator

“…Plasmids-Plasmids containing GFP-Myosin VI globular tail (GFPMyoVI-Tail), wt-CFTR (CFTR), and GFP-wt-CFTR (GFP-CFTR) constructs were generated as described (41,51). The GFP-CFTR Y/A LL/AA construct was generated from GFP-CFTR by the following alanine substitutions that disrupt the tyrosine and dileucine endocytic motifs, Y1424A, L1430A, and L1431A, using site-directed mutagenesis as described (52).…”

The MRP family of drug efflux pumps

“…However, two other reports point to the possibility of C-terminal determinants. Apical routing of CFTR involves an interaction with a PDZ scaffolding protein that is mediated by a PDZ interaction motif at the C-terminus of CFTR (Moyer et al, 1999). Recently, MRP2 was also determined to have a functional PDZ interaction motif ('TKF') at its extreme C-terminus, as indicated by yeast two-hybrid and in vitro overlay assays (Kocher et al, 1999;Hegedus et al, 2003), and deletion or mutation of this sequence in MRP2-GFP was reported to confer basolateral localization in transfected MDCK cells (Harris et al, 2001).…”