Inactivation of metal-induced metallothionein gene expression by protein kinase C inhibitor

Abstract

The involvement of protein kinase C (PKC) in the induction of metallothionein (MT) gene expression by non-metallic compounds has been reported. However, whether PKC participates in the metal-induced MT gene transcription remains unclear. We used PKC inhibitor, H7 and cherelythrine, to treat CHO CdR and GH3 cells, and found that both Cd- and Zn-induced MT gene expressions were blocked. Protein kinase A (PKA) was apparently not involved in the induction since PKA inhibitor, HA1004, did not affect the expression of MT gene. By using constructs with a reporter gene linked to different regions of MT promoter, we observed that transcription factor API was not associated with the induction. The inactivation of MT gene expression by PKC inhibitor was not due to block of Cd transport since cellular Cd content was not affected by the inhibitor. However, the PKC inhibitor dramatically reduced cellular Zn accumulation when stimulated by Zn ions. Further analyses of MT transcriptional factor, MTF-1, by semi-quantitative reverse transcriptase-polymerase chain reaction indicated that MTF-1 gene expression was not changed by the PKC inhibitor. We also found that vanadate, a phosphatase inhibitor, increased both basal and induced level of MT gene expression. These results suggest that MT gene expression induced by metal ions involves a PKC mediated phosphorylation pathway.