Heat shock protein 27, one of the low-molecular-weight stress proteins, is recognized as a molecular chaperone, however, other functions have not yet been well established. Phosphorylated heat shock protein 27 levels inversely correlate with the progression of human hepatocellular carcinoma. This study shows that phosphorylated heat shock protein 27 interferes with cell growth of the hepatocellular carcinoma derived HuH7 cells in the presence of the proinflammatory cytokine, tumor necrosis factor-a, via inhibition of the sustained activation of the extracellular-signal-regulated kinase signal pathway. The activities of Raf/ extracellular-signal- regulated kinase and subsequent activator protein-1 transactivation, and the induction levels of cyclin D1 were lower in HuH7 cells transfected with phosphorylated heat shock protein 27 than those with unphosphorylated heat shock protein 27. Moreover, phosphorylated heat shock protein 27 up-regulated the levels of p38 mitogen-activated protein kinase and mitogen-activated protein kinase phosphatase-1, an inhibitory protein of extracellular-signal-regulated kinase. These results indicate that phosphorylated heat shock protein 27 might suppress the extracellular-signal-regulated kinase activity in the hepatocellular carcinoma cells via two separate pathways in an inflammatory state. The extracellular-signal-regulated kinase activity is inversely correlated with phosphorylated heat shock protein 27 at serine-15, also in human hepatocellular carcinoma tissues in vivo. Since, the extracellular-signal-regulated kinase signal pathway is a major proliferation signal of hepatocellular carcinoma, and activator protein-1 activation is an early event in hepatocarcinogenesis. These findings strongly suggest that the control of the phosphorylated heat shock protein 27 levels could be a new therapeutic strategy especially to counter the recurrence of hepatocellular carcinoma.