MKK4 is a novel target for the inhibition of tumor necrosis factor-alpha-induced vascular endothelial growth factor expression by myricetin.
Tumor necrosis factor-alpha (TNF-alpha) is a mediator of multiple inflammatory diseases. Vascular endothelial growth factor (VEGF) plays a critical role in TNF-alpha-mediated diseases. We investigated the inhibitory effects of 3,3',4',5,5',7-hexahydroxyflavone (myricetin), an abundant natural flavonoid, on TNF-alpha-induced VEGF upregulation and the underlying molecular mechanism. Myricetin is a direct inhibitor of mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinase 1 (MEK1) and inhibits neoplastic cell transformation. We found that myricetin inhibited TNF-alpha-induced VEGF expression in JB6 P+ mouse epidermal cells by targeting MAPK kinase 4 (MKK4), as well as MEK1. The activation of activator protein-1 by TNF-alpha was inhibited by myricetin in a dose-dependent manner. The phosphorylation of c-Jun N-terminal kinase (JNK) and ERK was inhibited by myricetin, but not the phosphorylation of their upstream kinases MKK4 and MEK1. TNF-alpha-induced VEGF expression was inhibited by SP600125 and U0126, which are inhibitors of JNK and MEK, respectively. Myricetin inhibited TNF-alpha-induced MKK4 activity and bound glutathione S-transferase-MKK4 directly by competing with ATP. Computer modeling suggested that myricetin docks onto the ATP-binding site in MKK4, which is located between the N- and C-lobes of the kinase domain. Overall, our results indicate that myricetin has potent chemopreventive effects against TNF-alpha-related disease, mainly by targeting MKK4 and MEK1.
Kim
Jong-Eun
JE
Department of Agricultural Biotechnology and Research Institute for Agriculture and Life Sciences, Seoul National University, San 56-1, Shillim-dong, Gwanak-gu, Seoul 151-921, Republic of Korea.
Kwon
Jung Yeon
JY
Lee
Dong Eun
DE
Kang
Nam Joo
NJ
Heo
Yong-Seok
YS
Lee
Ki Won
KW
Lee
Hyong Joo
HJ
D000818:Animals; D015153:Blotting, Western; D004797:Enzyme-Linked Immunosorbent Assay; D005419:Flavonoids / Q000494:pharmacology*; D047468:Immunoprecipitation; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D008958:Models, Molecular; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors*; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis* / Q000378:metabolism
D005419:Flavonoids; D014409:Tumor Necrosis Factor-alpha; D042461:Vascular Endothelial Growth Factor A; C040015:myricetin; D020928:Mitogen-Activated Protein Kinases