A novel enhancement of SV40 enhancer activity by treatment of mouse neuroblastoma N18TG2 with protein kinase inhibitor H-7.

Abstract

To determine the role of protein phosphorylation in transcription regulation, we have treated mouse neuroblastoma N18TG2 cells with the protein kinase inhibitor H-7 and tested its effect on transcription. After the preculture and transfection in the presence of H-7, the cell preparation was divided in half and cultured with and without H-7. The level of CAT expression of pSV2-CAT was found to be higher in the cells cultured in the absence of H-7 than in those cultured in the presence of H-7. This difference was observed only after pretreatment of the cells with H-7, suggesting that withdrawal of H-7 from the culture medium after preculture with H-7 gave an enhancing effect on CAT expression. This phenomenon was also observed with transformants that expressed the CAT gene of pSV2-CAT stably. The 72 base-pair (bp) repeat of SV40 DNA was responsible for this difference in CAT expression. A similar effect of H-7 on the SV40 enhancer activity was observed in mouse neuroblastoma x rat glioma hybrid NG108-15 cells, but not in rat glioma C6-BU-1 cells.