Cross-talk between protein kinases Cζ and B in cyclic AMP-mediated sodium taurocholate co-transporting polypeptide translocation in hepatocytes

Abstract

Cyclic AMP stimulates taurocholate (TC) uptake and sodium taurocholate co-transporting polypeptide (Ntcp) translocation in hepatocytes via the phosphoinositide-3 kinase (PI3K) signaling pathway. The aim of the present study was to determine whether protein kinase (PK) Cζ, one of the downstream mediators of the PI3K signaling pathway, is involved in cAMP-mediated stimulation of TC uptake. Studies were conducted in isolated rat hepatocytes and in HuH-7 cells stably transfected with rat liver Ntcp (HuH-Ntcp cells). Studies in hepatocytes showed that cAMP activates PKCζ in a PI3K-dependent manner without inducing translocation of PKCζ to the plasma membrane. Inhibition of cAMP-induced PKCζ activity by myristoylated PKC (ζ/λ) pseudosubstrate, a specific inhibitor of PKCζ, and Gö 6850, a PKC inhibitor, resulted in inhibition of cAMP-induced increases in TC uptake and Ntcp translocation. Studies in HuH-Ntcp cells showed that inhibition of cAMP-induced PKCζ activation by dominant-negative (DN) PKCζ resulted in inhibition of cAMP-induced increases in TC uptake and Ntcp translocation. DN PKCζ also inhibited wild-type PKCζ-induced increases in PKCζ activity, TC uptake, and Ntcp translocation. Myristoylated PKC (ζ/λ) pseudosubstrate and DN PKCζ also inhibited cAMP-induced activation of PKB in hepatocytes and HuH-Ntcp cells, respectively. Neither DN PKB nor constitutively active PKB affected cAMP-induced activation of PKCζ, and wild-type PKCζ did not activate PKB. Taken together, these results suggest that cAMP-induced activation of PKB is dependent on cAMP-induced stimulation of PKCζ. It is proposed that cAMP-induced Ntcp translocation involves the activation of the PI3K/PKCζ signaling pathway followed by the activation of the PI3K/PKB signaling pathway.