Activation of 3-Phosphoinositide-dependent Kinase 1 (PDK1) and Serum- and Glucocorticoid-induced Protein Kinase 1 (SGK1) by Short-chain Sphingolipid C4-ceramide Rescues the Trafficking Defect of ΔF508-Cystic Fibrosis Transmembrane Conductance Regulator (ΔF508-CFTR)

Cystic fibrosis (CF) is due to a folding defect in the CF transmembrane conductance regulator (CFTR) protein. The most common mutation, ΔF508, prevents CFTR from trafficking to the apical plasma membrane. Here we show that activation of the PDK1/SGK1 signaling pathway with C4-ceramide (C4-CER), a non-toxic small molecule, functionally corrects the trafficking defect in both cultured CF cells and primary epithelial cell explants from CF patients. The mechanism of C4-CER action involves a series of mutual autophosphorylation and phosphorylation events between PDK1 and SGK1. Detailed mechanistic studies indicate that C4-CER initially induces autophosphorylation of SGK1 at Ser 422 . SGK1[Ser(P) 422 ] and C4-CER coincidently bind PDK1 and permit PDK1 to autophosphorylate at Ser 241 . Then PDK1[Ser(P) 241 ] phosphorylates SGK1[Ser(P) 422 ] at Thr 256 to generate fully activated SGK1[Ser 422 , Thr(P) 256 ]. SGK1[Ser(P) 422 ,Thr(P) 256 ] phosphorylates and inactivates the E3 ubiquitin ligase Nedd4-2. ΔF508-CFTR is thus free to traffic to the plasma membrane. Importantly, C4-CER-mediated activation of both PDK1 and SGK1 is independent of the PI3K/Akt/mammalian target of rapamycin signaling pathway. Physiologically, C4-CER significantly increases maturation and stability of ΔF508-CFTR ( t ½ ∼10 h), enhances cAMP-activated chloride secretion, and suppresses hypersecretion of interleukin-8 (IL-8). We suggest that candidate drugs for CF directed against the PDK1/SGK1 signaling pathway, such as C4-CER, provide a novel therapeutic strategy for a life-limiting disorder that affects one child, on average, each day.The discovery of small molecules to correct the trafficking defect of the mutant ΔF508-CFTR protein has been challenging. Results C4-ceramide rescues and stabilizes ΔF508-CFTR. High basal secretion of interleukin-8 is also suppressed. Conclusion Results identify a novel mechanism by which C4-ceramide activates the PDK1/SGK1 pathway, thereby rescuing ΔF508-CFTR. Significance C4-ceramide may be a novel candidate therapeutic for CF patients.