Dibutyryl cAMP (DbcAMP) Induces Serum and Glucocorticoid‐Induced Kinase1 (sgk1) Expression and Initiates Amiloride‐Sensitive Na+ Transport in Rat Submandibular Gland Epithelial (SMG‐C6) Cells

Abstract

Transepithelial Na+ transport occurs via Na+channels (ENaC) which consist of three protein subunits, α, β, and γ . SMG-C6 cells cultured in the absence of hydrocortisone or DbcAMP do not express sgk1 or possess Na+ transport properties. Treatment of SMG-C6 cells with DbcAMP for 24h up-regulated α-ENaC mRNA and protein expression and amiloride-sensitive Na+ transport but did not affect endogenous β-or γ-ENaC protein levels. We hypothesized that the initiation of α-ENaC expression and function by DbcAMP is preceded by an endogenous increase in sgk1. Treatment of SMG-C6 cells with DbcAMP increased sgk1 protein expression after 2h which remained elevated for 24h. DbcAMP-induced sgk1 mRNA expression was decreased by actinomycin D and by inhibitors of PKA (H-89 and KT5720). Inhibition of the PI3-kinase pathway with LY294002 or dominant negative PI3-kinase reduced not only the phosphorylation of CREB and sgk1 mRNA but also after 24h, Na+ transport. A combination of either PKA inhibitor with dominant negative PI3-kinase synergistically abolished Na+ transport after 24h. The addition of LY294002 to cells previously exposed to DbcAMP for 24h significantly reduced DbcAMP-induced Na+ transport. Thus, sgk1is induced by DbcAMP via PI3-kinase and PKA signaling pathways prior to elevated α-ENaC expression and function. We speculate that in SMG-C6 cells, maintenance of DbcAMP–induced Na+ transport is dependent upon the activation of sgk1.