Chlorpromazine activates p21Waf1/Cip1 gene transcription via early growth response-1 (Egr-1) in C6 glioma cells.
2-Chloro-10-[3(-dimethylamino)propyl]phenothiazinemonohydrochloride (chlorpromazine) is a phenothiazine derivative used clinically to control psychotic disorders. It also exhibits an anticancer activity. Treatment with chlorpromazine (CPZ) results in cell-cycle arrest at the G2/M phase in rat C6 glioma cells. CPZ reduces the expression of cell cycle-related proteins, such as cyclin D1, cyclin A, and cyclin B1, but causes an increase in the p21(Waf1/Cip1) level. The molecular mechanism by which CPZ regulates p21(Waf1/Cip1) expression is unknown. Here, we provide evidence that CPZ activates the p21(Waf1/Cip1) gene promoter via induction of the transcription factor early growth response-1 (Egr-1) independently of p53 in C6 cells. A point mutation in the Egr-1-binding motif within the p21(Waf1/Cip1) promoter abrogated promoter inducibility due to CPZ. Forced expression of Egr-1 enhanced p21(Waf1/Cip1) promoter activity. In contrast, knockdown of endogenous Egr-1 by small interference RNA attenuated CPZ-induced p21(Waf1/Cip1) promoter activity. A chromatin immunoprecipitation assay demonstrated that Egr-1 binds to the p21(Waf1/Cip1) gene promoter. Further analysis showed that the ERK and JNK MAP kinases are required for induction of Egr-1 by CPZ. Finally, stable silencing of Egr-1 expression lead to attenuated CPZ-inducible p21(Waf1/Cip1) expression and inhibited G2/M phase cell-cycle arrest. These results demonstrate that a functional link between ERK and JNK MAP kinase pathways and p21(Waf1/Cip1) induction via Egr-1 contributes to CPZ-induced anticancer activity in C6 glioma cells.
Shin
Soon Young
SY
Institute of Biomedical Science and Technology, Konkuk University Hospital, Seoul 143-729, Korea.
Kim
Chang Gun
CG
Kim
Se Hyun
SH
Kim
Yong Sik
YS
Lim
Yoongho
Y
Lee
Young Han
YH
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects; D002746:Chlorpromazine / Q000494:pharmacology*; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000378:metabolism*; D051766:Early Growth Response Protein 1 / Q000235:genetics / Q000378:metabolism*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D005910:Glioma / Q000188:drug therapy / Q000378:metabolism*; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D034741:RNA, Small Interfering; D051381:Rats; D014158:Transcription, Genetic / Q000187:drug effects; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism
D000970:Antineoplastic Agents; D050759:Cyclin-Dependent Kinase Inhibitor p21; C497325:EGR1 protein, human; D051766:Early Growth Response Protein 1; D034741:RNA, Small Interfering; D016159:Tumor Suppressor Protein p53; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D002746:Chlorpromazine