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 &x25b5;F508-cystic fibrosis transmembrane conductance regulator (&x25b5;F508-CFTR)

Cystic fibrosis (CF) is due to a folding defect in the CF transmembrane conductance regulator (CFTR) protein. The most common mutation, &x25B5;F508, 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 Ser⁴²². SGK1[Ser(P)⁴²² and C4-CER coincidently bindPDK1and permitPDK1to autophosphorylate at Ser²⁴¹. Then PDK1[Ser(P)sup>241] phosphorylates SGK1[Ser(P)⁴²²] at Thr²⁵⁶ to generate fully activated SGK1[Ser⁴²², Thr(P)²⁵⁶]. SGK1[Ser(P)⁴²²,Thr(P)²⁵⁶] phosphorylates and inactivates the E3 ubiquitin ligase Nedd4-2. &x25B5;F508-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 &x25B5;F508-CFTR (t_1/2∼10 h), enhances cAMP-activated chloride secretion, and suppresses hypersecretion of interleukin-8 (IL-8).Wesuggest 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.