15-Deoxy-Delta(12,14) prostaglandin J2 (15d-PGJ2) is a natural ligand for the peroxisome proliferator-activated receptor gamma (PPARgamma) that exhibits antiproliferative activity in colon cancer cells, but its mechanism of action is still poorly understood. In this study, we showed that Krüppel-like factor 4 (KLF4) is one of the downstream effectors of 15d-PGJ2. Treatment of HT-29 cells with 15d-PGJ2 resulted in up-regulation of both KLF4 mRNA and protein expression, and these increases were also observed in other colon cancer cell lines. Down-regulation of KLF4 expression by small interfering RNA (siRNA) targeting KLF4 reduced 15d-PGJ2-mediated G1 phase arrest, suggesting that KLF4-mediated function of 15d-PGJ2. The effect of 15d-PGJ2 on KLF4 expression seems not to involve its nuclear receptor PPARgamma, in that our data show that:1) KLF4 gene promoter does not contain putative PPRE sequence, 2) 15d-PGJ2 rapidly activates extracellular signal-regulated kinase (ERK) and induces KLF4 mRNA expression, 3) KLF4 is induced by 15d-PGJ2 but not by rosiglitazone, a synthetic PPARgamma ligand, and 4) 15d-PGJ2 is unable to stimulate PPAR-dependent promoter activity in the absence of cotransfected PPARgamma. Moreover, 15d-PGJ2-mediated KLF4 mRNA expression was blocked by 2'-amino-3'-methoxyflavone (PD98059) or 1,4-diamino-2,3-dicyano-1,4-bis(methylthio)butadiene (U0126), two ERK kinase MAP inhibitors, whereas the phosphoinositol-3 kinase inhibitors wortmannin and 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one (LY294002) had no such effect. Furthermore, KLF4 induction by 15d-PGJ2 occurred only in signal transducer and activator of transcription 1 (STAT1)-expressing, not in STAT1-knockout cells. Together, these results suggest that 15d-PGJ2-induced growth inhibition of colon cancer cells is mediated, at least in part, through up-regulation of KLF4 expression. This induction is unlikely to be mediated through the PPARgamma receptor but may involve the mitogen-activated protein kinase kinase/ERK pathway and is STAT1-dependent.