Inhibition of the ERK phosphorylation plays a role in terbinafine-induced p21 up-regulation and DNA synthesis inhibition in human vascular endothelial cells.
Previously, we showed that terbinafine (TB) induces cell-cycle arrest in cultured human umbilical vein endothelial cells (HUVEC) through an up-regulation of the p21 protein. The aim of this study is to delineate the molecular mechanisms underlying TB-induced increase of p21 protein. RT-PCR analysis demonstrated that the mRNA levels of p21 and p53 were increased in the TB-treated HUVEC. The p21 promoter activity was also increased by TB treatment. Transfection of HUVEC with p53 dominant negative (DN) abolished the TB-induced increases of p21 promoter activity and protein level, suggesting that the TB-induced increase of p21 is p53-dependent. Western blot analysis demonstrated that TB decreased the levels of phosphorylated extracellular signal-regulated kinase (ERK). Over-expression of mitogen-activated protein kinase (MEK)-1, the immediate upstream activator kinase of ERK, abolished the TB-induced increases of p21 and p53 protein and decrease of thymidine incorporation. The ERK inhibitor (PD98059) enhanced the TB-induced inhibition of thymidine incorporation into HUVEC. Taken together, these data suggest that the decrease of ERK activity plays a role in the TB-induced up-regulation of p21 in HUVEC. On the other hand, pretreatment of the cells with geranylgeraniol (GGOH), farnesol (FOH), or Ras inhibitor peptide did not affect the TB-induced decrease of thymidine incorporation. Taken together, our results suggest that TB might cause a decrease of MEK, which in turn up-regulates p53 through the inhibition of ERK phosphorylation, and finally causes an increase of p21 expression and cell-cycle arrest.
Ho
Pei-Yin
PY
Graduate Institute of Cell and Molecular Biology, Taipei Medical University, Taipei, Taiwan.
Hsu
Sung-Po
SP
Liang
Yu-Chih
YC
Kuo
Min-Liang
ML
Ho
Yuan-Soon
YS
Lee
Wen-Sen
WS
D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000187:drug effects / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis / Q000187:drug effects*; D004730:Endothelium, Vascular / Q000187:drug effects* / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D019950:Mitogen-Activated Protein Kinase 1 / Q000187:drug effects / Q000378:metabolism; D009281:Naphthalenes / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000077291:Terbinafine; D013936:Thymidine / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000187:drug effects / Q000378:metabolism; D014471:Umbilical Veins / Q000187:drug effects / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects*
D050759:Cyclin-Dependent Kinase Inhibitor p21; D004791:Enzyme Inhibitors; D009281:Naphthalenes; D016159:Tumor Suppressor Protein p53; D004247:DNA; D048049:Extracellular Signal-Regulated MAP Kinases; D019950:Mitogen-Activated Protein Kinase 1; D000077291:Terbinafine; D013936:Thymidine