The regulation of CFTR by protein-protein interactions Public Deposited

Downloadable Content

Download PDF
Last Modified
  • March 22, 2019
  • Thelin, William R.
    • Affiliation: School of Medicine, Department of Cell Biology and Physiology
  • Cystic fibrosis (CF) is an autosomal recessive disease resulting from the misregulation of epithelial ion transport. CF is caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), an apical membrane chloride channel expressed in polarized epithelial cells. To identify factors that regulate CFTR activity, we utilized biochemical and proteomics approaches to identify novel CFTR binding proteins. We find that the C-terminus of CFTR directly interacts with the serine/threonine phosphatase PP2A. PP2A is a heterotrimeric phosphatase composed of a catalytic subunit and two divergent regulatory subunits (A and B), which mediate the cellular localization and substrate specificity of the enzyme. By mass spectrometry, we identified the exact PP2A regulatory subunits associated with CFTR as Aα and B’ε, and find that the B’ε subunit binds CFTR directly. PP2A subunits localize to the apical surface of airway epithelia and PP2A phosphatase activity co-purifies with CFTR in Calu-3 cells. In functional assays, PP2A inhibition blocks the rundown of basal CFTR currents and increases channel activity in excised patches of airway epithelia and in intact mouse jejunum. Moreover, PP2A inhibition increases the pericilliary liquid in cultures of well differentiated human bronchial epithelial cells by a CFTR-dependent mechanism. Thus, PP2A is a relevant CFTR phosphatase in epithelial tissues and may be a clinically relevant drug target for CF. Additionally, the N-terminus of CFTR directly interacts with two actin binding proteins, filamin A and filamin B. In polarized epithelial cells, filamins are highly localized to the sub-apical compartment where they likely interact with CFTR at or near the plasma membrane. We find that CFTR and filamins specifically interact by co-immunoprecipitation and that a disease-causing mutation in CFTR, serine 13 to phenylalanine (S13F), disrupts this interaction. Consistent with the loss of cytoskeletal anchorage, S13F CFTR displays decreased cell surface levels and less confinement at the plasma membrane relative to wildtype CFTR. Furthermore, S13F CFTR is more rapidly degraded compared to wild-type CFTR which correlates with the accumulation of S13F CFTR in the lysosomes. Taken together, these data suggest the filamins regulate the cell surface stability and endocytic trafficking of CFTR.
Date of publication
Resource type
Rights statement
  • In Copyright
  • Milgram, Sharon L.
Degree granting institution
  • University of North Carolina at Chapel Hill
  • Open access

This work has no parents.