Identification of Two Nickel Ion-Induced Genes, NCI16 and Pc GST1 , in Paramecium caudatum

Abstract

ABSTRACT Here, we describe the isolation of two nickel-induced genes in Paramecium caudatum , NCI16 and Pc GST1 , by subtractive hybridization. NCI16 encoded a predicted four-transmembrane domain protein (∼16 kDa) of unknown function, and Pc GST1 encoded glutathione S -transferase (GST; ∼25 kDa) with GST and glutathione peroxidase (GPx) activities. Exposing cells to cobalt chloride also caused the moderate upregulation of NCI16 and Pc GST1 mRNAs. Both nickel sulfate and cobalt chloride dose dependently induced NCI16 and Pc GST1 mRNAs, but with different profiles. Nickel treatment caused a continuous increase in Pc GST1 and NCI16 mRNA levels for up to 3 and 6 days, respectively, and a notable increase in H 2 O 2 concentrations in P. caudatum . NCI16 expression was significantly enhanced by incubating cells with H 2 O 2 , implying that NCI16 induction in the presence of nickel ions is caused by reactive oxygen species (ROS). On the other hand, Pc GST1 was highly induced by the antioxidant tert -butylhydroquinone (tBHQ) but not by H 2 O 2 , suggesting that different mechanisms mediate the induction of NCI16 and Pc GST1 . We introduced a luciferase reporter vector with an ∼0.42-kb putative Pc GST1 promoter into cells and then exposed the transformants to nickel sulfate. This resulted in significant luciferase upregulation, indicating that the putative Pc GST1 promoter contains a nickel-responsive element. Our nickel-inducible system also may be applicable to the efficient expression of proteins that are toxic to host cells or require temporal control.