In this study, we demonstrated a mechanism underlying fisetin-induced apoptosis. Fisetin markedly induced apoptosis via induction of DR5 expression. In addition, fisetin induced up-regulation of p53 expression at the post-translational level, and down-regulation of p53 by siRNA-inhibited up-regulation of DR5 expression in fisetin-treated cells. Therefore, our results suggest that fisetin induces apoptosis though p53-mediated up-regulation of DR5 expression.
Several papers have reported that fisetin induced apoptosis in multiple cancer cell lines, and in addition suggested the molecular mechanisms related with induction of apoptosis [7
]. First, fisetin modulates the MAPK signaling pathway. Fisetin induced apoptosis in human non-small lung cancer cells and cervical cancer cells through activation of the MAPK signaling pathway [7
]. Sustained activation of ERK and ERK-mediated CHOP expression is associated with fisetin-induced apoptosis [7
]. Second, fisetin inhibited NF-κB signaling pathways. Fisetin induced apoptosis in colon cancer cells as well as in bladder cancer cells through inhibition of NF-κB signaling pathways [26
]. Inhibition of NF-κB signaling resulted in down-regulation of anti-apoptotic proteins (c-FLIP and cyclooxygenase-2). Third, p53 plays critical roles on fisetin-induced apoptosis. Two previous studies reported that induction of p53 expression contributed to induction of apoptosis in human colon cancer cells [22
] and bladder cancer cells [27
]. In contrast, fisetin can also induce apoptosis in a p53-independent manner. In gastric cancer cells, although fisetin increased p53 protein expression, inhibitor of p53 did not reduce fisetin-mediated apoptosis [3
]. Therefore, roles of p53 on fisetin-induced apoptosis are different depending on cell type. Finally, ROS are important signaling molecules on fisetin-induced apoptosis. Production of ROS by fisetin treatment is associated with induction of apoptosis in multiple myeloma [24
] and oral cancer cells [8
]. An ROS scavenger molecule, NAC, reduced fisetin-induced apoptosis in both cells. Recently, Wu et al. suggested that NAC enhanced fisetin-induced apoptosis through inhibition of ERK phosphorylation in colon cancer cells [28
]. In this study, NAC completely inhibited H2
-induced ROS production and apoptosis, whereas fisetin-induced ROS production did not alter with NAC treatment [28
]. Lim et al. suggested that the enhanced effect of NAC on fisetin-mediated apoptosis is independent of the scavenging effect of NAC. In our study, NAC did not enhance fisetin-induced apoptosis, and the ROS scavenger, GEE, also had no effect on apoptosis (Figure 6
B). Therefore, ROS signaling is not associated with fisetin-induced apoptosis in human renal carcinoma Caki cells.
As shown in Figure 2
C and Figure 3
C, fisetin induced DR5 expression and subsequently increased DR5 presentation on cell surface. Previous studies reported that death receptors could activate in a ligand-independent manner [29
]. For example, palmitate increased ER stress-induced CHOP expression, resulting in the upreguatlion of DR5 in hepatocytes [29
]. Although human recombinant DR5-Fc chimera proteins inhibited TRAIL-induced apoptosis, plamitate-induced apoptosis through induction of DR5 was not blocked by DR5-Fc [29
]. In addtion, persistent ER stress-mediated up-regulation of DR5 expression also promoted apoptosis through ligand-independent DR5 activation [30
]. Furthermore, Lim et al. reported that the recruitment of DR5 to a lipid raft induced DR5 activation, followed by apoptosis in ursodeoxycholic acid-treated human gastric cancer cells [31
]. Therefore, this strategy to induce up-regulation of DR5 might be effective to promote cancer cell death. Taken together, our data suggested that fisetin induced apoptosis through p53-mediated up-regulation of DR5 expression in human renal carcinoma Caki cells, and these findings provide an important molecular mechanism involved in fisetin-induced apoptosis.