Regulation of TNF-α-activated PKC-ζ signaling by the human biliverdin reductase:: identification of activating and inhibitory domains of the reductase

Human biliverdin reductase ( hBVR) is a dual function enzyme: a catalyst for bilirubin formation and a S/T/Y kinase that shares activators with protein kinase C (PKC)-zeta, including cytokines, insulin, and reactive oxygen species ( ROS). Presently, we show that hBVR increases PKC-zeta autophosphorylation, stimulation by TNF-alpha, as well as cytokine stimulation of NF-kappa B DNA binding and promoter activity. S-149 in hBVR S/T kinase domain and S-230 in (YLSF)-F-230 in hBVR's docking site for the SH2 domain of signaling proteins are phosphorylation targets of PKC-zeta. Two hBVR- based peptides, (KRNRYLSF)-F-230 (# 1) and KKRILHC281 (# 2), but not their S -> A or C -> A derivatives, respectively, blocked PKC-zeta stimulation by TNF-alpha and its membrane translocation. The C- terminal- based peptide KYCCSRK296 (# 3), enhanced PKC-zeta stimulation by TNF-alpha; for this, Lys(296) was essential. In metabolically P-32-labeled HEK293 cells transfected with hBVR or PKC-zeta, TNF-alpha increased hBVR phosphorylation. TNF-alpha did not stimulate PKC-zeta in cells infected with small interfering RNA for hBVR or transfected with hBVR with a point mutation in the nucleotide-binding loop (G(17)), S149, or S230; this was similar to the response of kinase-dead PKC-zeta(K281R). We suggest peptide #1 blocks PKC-zeta-docking site interaction, peptide #2 disrupts function of the PKC-zeta C1 domain, and peptide # 3 alters ATP presentation to the kinase. The findings are of potential significance for development of modulators of PKC-zeta activity and cellular response to cytokines.