NHE-RF1 protein rescues ΔF508-CFTR function

In cystic fibrosis ( CF), the Delta F508- CFTR anterograde trafficking from the endoplasmic reticulum to the plasma membrane is inefficient. New strategies for increasing the delivery of Delta F508- CFTR to the apical membranes are thus pathophysiologically relevant targets to study for CF treatment. Recent studies have demonstrated that PDZ-containing proteins play an essential role in determining polarized plasma membrane expression of ionic transporters. In the present study we have hypothesized that the PDZ- containing protein NHE-RF1, which binds to the carboxy terminus of CFTR, rescues Delta F508- CFTR expression in the apical membrane of epithelial cells. The plasmids encoding Delta F508- CFTR and NHE- RF1 were intranuclearly injected in A549 or Madin- Darby canine kidney ( MDCK) cells, and Delta F508- CFTR channel activity was functionally assayed using SPQ fluorescent probe. Cells injected with Delta F508- CFTR alone presented a low chloride channel activity, whereas its coexpression with NHE- RF1 significantly increased both the basal and forskolin- activated chloride conductances. This last effect was lost with Delta F508- CFTR deleted of its 13 last amino acids or by injection of a specific NHE- RF1 antisense oligonucleotide, but not by NHE- RF1 sense oligonucleotide. Immunocytochemical analysis performed in MDCK cells transiently transfected with Delta F508- CFTR further revealed that NHE- RF1 specifically determined the apical plasma membrane expression of Delta F508- CFTR but not that of a trafficking defective mutant potassium channel ( KCNQ1). These data demonstrate that the modulation of the expression level of CFTR protein partners, like NHE- RF1, can rescue Delta F508- CFTR activity.