LMTK2-mediated Phosphorylation Regulates CFTR Endocytosis in Human Airway Epithelial Cells

• Published in: The Journal of biological chemistry Vol. 289; no. 21; pp. 15080 - 15093
• Main Authors: Luz, Simão, Cihil, Kristine M., Brautigan, David L., Amaral, Margarida D., Farinha, Carlos M., Swiatecka-Urban, Agnieszka
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 23-05-2014; American Society for Biochemistry and Molecular Biology
• Subjects: Cell Biology; Cell Line; Cell Line, Tumor; Cell Membrane - metabolism; Cells, Cultured; CFTR; Chloride Transport; Chlorides - metabolism; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator - genetics; Cystic Fibrosis Transmembrane Conductance Regulator - metabolism; Endocytosis; Epithelial Cell; Epithelial Cells - metabolism; HEK293 Cells; Humans; Immunoblotting; Immunoprecipitation; Membrane Proteins - genetics; Membrane Proteins - metabolism; Mutation; Phosphorylation; Protein Phosphorylation; Protein-Serine-Threonine Kinases - genetics; Protein-Serine-Threonine Kinases - metabolism; Respiratory System - cytology; RNA Interference; Serine - genetics; Serine - metabolism; Epithelial Cell; Chloride Transport; Endocytosis; Protein Phosphorylation; Cystic Fibrosis; CFTR
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M114.563742

Cystic fibrosis transmembrane conductance regulator (CFTR) is a Cl−-selective ion channel expressed in fluid-transporting epithelia. Lemur tyrosine kinase 2 (LMTK2) is a transmembrane protein with serine and threonine but not tyrosine kinase activity. Previous work identified CFTR as an in vitro substrate of LMTK2, suggesting a functional link. Here we demonstrate that LMTK2 co-immunoprecipitates with CFTR and phosphorylates CFTR-Ser737 in human airway epithelial cells. LMTK2 knockdown or expression of inactive LMTK2 kinase domain increases cell surface density of CFTR by attenuating its endocytosis in human airway epithelial cells. Moreover, LMTK2 knockdown increases Cl− secretion mediated by the wild-type and rescued ΔF508-CFTR. Compared with the wild-type CFTR, the phosphorylation-deficient mutant CFTR-S737A shows increased cell surface density and decreased endocytosis. These results demonstrate a novel mechanism of the phospho-dependent inhibitory effect of CFTR-Ser737 mediated by LMTK2 via endocytosis and inhibition of the cell surface density of CFTR Cl− channels. These data indicate that targeting LMTK2 may increase the cell surface density of CFTR Cl− channels and improve stability of pharmacologically rescued ΔF508-CFTR in patients with cystic fibrosis.
The CFTR-Ser737 site has a phospho-dependent inhibitory effect on Cl− secretion.
LMTK2 phosphorylation of CFTR-Ser737 facilitates endocytosis, reduces cell surface density of CFTR, and inhibits Cl− secretion.
Targeting LMTK2 regulates the cell surface density of CFTR Cl− channels.
Targeting LMTK2 in CF patients may stabilize ΔF508-CFTR pharmacologically rescued to the cell surface.