Induction of Early Growth Response-1 Gene Expression by Calmodulin Antagonist Trifluoperazine through the Activation of Elk-1 in Human Fibrosarcoma HT1080 Cells

Soon Young Shin; Seong-Yong Kim; Jung-Hye Kim; Do Sik Min; Jesang Ko; Ung-Gu Kang; Yong Sik Kim; Taeg Kyu Kwon; Mi Young Han; Young Ho Kim; Young Han Lee

doi:10.1074/jbc.M009465200

Induction of Early Growth Response-1 Gene Expression by Calmodulin Antagonist Trifluoperazine through the Activation of Elk-1 in Human Fibrosarcoma HT1080 Cells*

From the ^‡Department of Biochemistry and Molecular Biology, College of Medicine, Yeungnam University, Daegu 705–717, the ^¶Department of Physiology, College of Medicine, The Catholic University of Korea, Seoul 137-701, the ^‖Immunomodulation Research Center, University of Ulsan, Ulsan 138–736, the ^**Department of Psychiatry, College of Medicine, Seoul National University, Seoul 151–742, the ^‡Department of Immunology, School of Medicine, Keimyung University, Daegu 700–712, the ^§Cell Biology Laboratory, Korea Research Institute of Bioscience and Biotechnology, P. O. Box 115, Yusung, Taejon 305–600, and the ^§§Department of Microbiology, Kyungpook National University, Daegu 702–701, South Korea

Abstract

The early growth response gene-1 (Egr-1) is a transcription factor that plays an important role in cell growth and differentiation. It has been known that Egr-1 expression is down-regulated in many types of tumor tissues, including human fibrosarcoma HT1080 cells, and introduction of the Egr-1gene into HT1080 cells inhibits cell growth and tumorigenic potential. Trifluoperazine (TFP), a phenothiazine class calmodulin antagonist, is known to inhibit DNA synthesis and cell proliferation and potentially important in antitumor activities. To understand the regulatory mechanism of Egr-1, we investigated the effect of TFP on expression of Egr-1 in HT1080 cells. Herein, we report thatEgr-1 expression was increased by TFP in synergy with serum at the transcriptional level. Both the Ca²⁺/calmodulin-dependent protein kinase II inhibitor KN62 and the calcineurin inhibitor cyclosporin A enhanced TFP-dependent increase of Egr-1, suggesting that the Ca²⁺/calmodulindependent pathway plays a role in regulation of Egr-1 expression in HT1080 cells. The TFP-stimulated increase of the Egr-1 protein was preferentially inhibited by the MEK-specific inhibitor PD98059. In addition, activation of human Egr-1 promoter and the transcriptional activation of the ternary complex factor Elk-1 induced by TFP were inhibited both by pretreatment of PD98059 and by expression of the dominant-negative RasN17. These results indicate that the Ras/MEK/Erk/Elk-1 pathway is necessary for TFP-inducedEgr-1 expression. We propose that the calmodulin antagonist TFP stimulates Egr-1 gene expression by modulating Ras/MEK/Erk and activation of the Elk-1 pathway in human fibrosarcoma HT1080 cells.