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,
22,
24,
25,
26,
27,
28,
29]. 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,
25]. Sustained activation of ERK and ERK-mediated CHOP expression is associated with fisetin-induced apoptosis [
7,
25]. 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,
27]. 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 H
2O
2-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 6B). Therefore, ROS signaling is not associated with fisetin-induced apoptosis in human renal carcinoma Caki cells.
As shown in
Figure 2C and
Figure 3C, 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,
30,
31]. 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.