c-Cbl, CHIP or Nedd4-2 depletion do not affect rΔF508 CFTR endocytosis.

<p>CFBE41o-ΔF cells were transfected with control or c-Cbl, CHIP, or Nedd4-2 siRNA oligonucleotides. At 24 h after transfection, cells were transferred to Transwell filters and cultured for an additional 4–5 days. The last 24 h, the cells were cultured at 27°C to promote ΔF508 CFTR delivery to the cell surface. <b>(A)</b> Cell surface expression of rΔF508 CFTR after siRNA transfection of control (C), c-Cbl, CHIP or Nedd4-2 as indicated. The knockdown efficiency of c-Cbl, CHIP, and Nedd4-2 was >95%. β-actin was blotted as a loading control. <b>(B)</b> Representative blots showing the remaining surface rΔF508 CFTR after 2.5 min internalization at 37°C. <b>(C)</b> Quantitative analysis of rΔF508 CFTR internalization after 2.5 min warm-up following c-Cbl, CHIP, or Nedd4-2 siRNA depletion and low-temperature rescue. The rate of rΔF508 CFTR internalization was measured after 2.5 min warm-up as described in the Material and Methods section. Depletion of c-Cbl, CHIP, or Nedd4-2 had no significant (N.S.) effect on rΔF508 CFTR internalization (n = 3).