Recent evidence has revealed that senescence induction requires fine-tuned activation of p53, however, mechanisms underlying the regulation of p53 activity during senescence have not as yet been clearly established. We demonstrate here that SCF^Fbxo22-KDM4A is a senescence-associated E3 ligase targeting methylated p53 for degradation. We find that Fbxo22 is highly expressed in senescent cells in a p53-dependent manner, and that SCF^Fbxo22 ubiquitylated p53 and formed a complex with a lysine demethylase, KDM4A. Ectopic expression of a catalytic mutant of KDM4A stabilizes p53 and enhances p53 interaction with PHF20 in the presence of Fbxo22. SCF^Fbxo22-KDM4A is required for the induction of p16 and senescence-associated secretory phenotypes during the late phase of senescence. Fbxo22^−/− mice are almost half the size of Fbxo22^+/− mice owing to the accumulation of p53. These results indicate that SCF^Fbxo22-KDM4A is an E3 ubiquitin ligase that targets methylated p53 and regulates key senescent processes.

Luteolin is a natural flavonoid with strong anti–oxidative properties that is reported to have an anti–cancer effect in several malignancies other than bladder cancer. In this study, we describe the effect of luteolin on a human bladder cancer cell line, T24, in the context of the regulation of p21, thioredoxin‐1 (TRX1) and the mechanistic target of rapamycin (mTOR) pathway. Luteolin inhibited cell survival and induced G2/M cell‐cycle arrest, p21 upregulation and downregulation of phospho(p)‐S6, which is downstream of mTOR signaling. Luteolin also upregulated TRX1 and reduced intracellular reactive oxygen species production. In a subcutaneous xenograft mouse model using the rat bladder cancer cell line, BC31, tumor volumes were significantly decreased in mice orally administered luteolin compared to control. Immunohistochemical analysis revealed that increased p21 and decreased p‐S6 expression were induced in the luteolin treatment group. Moreover, in another in vivo N‐butyl‐N‐(4‐hydroxybutyl) nitrosamine (BBN)‐induced rat bladder cancer model, the oral administration of luteolin led to a trend of decreased bladder tumor dimension and significantly decreased the Ki67‐labeling index and p‐S6 expression. Furthermore, the major findings on the metabolism of luteolin suggest that both plasma and urine luteolin‐3ʹ‐O‐glucuronide concentrations are strongly associated with the inhibition of cell proliferation and mTOR signaling. Moreover, a significant decrease in the squamous differentiation of bladder cancer is attributed to plasma luteolin‐3ʹ‐glucuronide concentration. In conclusion, luteolin, and in particular its metabolized product, may represent another natural product‐derived therapeutic agent that acts against bladder cancer by upregulating p21 and inhibiting mTOR signaling.

The decision between survival and death in cells exposed to TNF relies on a highly regulated equilibrium between proapoptotic and antiapoptotic factors. The TNF-activated antiapoptotic response depends on several transcription factors, including NF-κB and its RelA/p65 subunit, that are activated through phosphorylation-mediated degradation of IκB inhibitors, a process controlled by the IκB kinase complex. Genetic studies in mice have identified the IκB kinase-related kinase TANK-binding kinase 1 (TBK1; also called NAK or T2K) as an additional regulatory molecule that promotes survival downstream of TNF, but the mechanism through which TBK1 exerts its survival function has remained elusive. Here we show that TBK1 triggers an antiapoptotic response by controlling a specific RelA/p65 phosphorylation event. TBK1-induced RelA phosphorylation results in inducible expression of plasminogen activator inhibitor-2 (PAI-2), a member of the serpin family with known antiapoptotic activity. PAI-2 limits caspase-3 activation through stabilization of transglutaminase 2 (TG2), which cross-links and inactivates procaspase-3. Importantly, Tg2 ^−/− mice were found to be more susceptible to apoptotic cell death in two models of TNF-dependent acute liver injury. Our results establish PAI-2 and TG2 as downstream mediators in the antiapoptotic response triggered upon TBK1 activation.

There is a need for exploration of new therapeutic strategies that target distinct molecular mechanisms of castration-resistant prostate cancer (CRPC) because its emergence following androgen deprivation therapy is a major clinical problem. In this report, we investigated the role of glutathione peroxidase 2 (GPX2) in CRPC. GPX2 expression was analyzed in rat and human CRPC cells. Next, we determined the proliferation rate and level of reactive oxygen species (ROS) in GPX2-small interfering RNA (siRNA)-transfected CRPC cells. For in vivo analysis, siRNA-transfected cells were subcutaneously implanted into normal and castrated nude mice. Further, immunohistochemical and prognostic analyses of GPX2 were performed using human specimens. Silencing of GPX2 caused significant growth inhibition and increased intracellular ROS in both rat (PCai1) and human (PC3) CRPC cells. Flow cytometry and western blot analyses revealed that the decrease in proliferation rate of the GPX2-silenced cells was due to cyclin B1-dependent G₂/M arrest. Furthermore, knockdown of Gpx2 inhibited tumor growth of PCai1 cells in castrated mice. Immunohistochemical analyses indicated that expression of GPX2 was significantly higher in residual cancer foci after neoadjuvant hormonal therapy than in hormone naive cancer foci. Moreover, patients with high GPX2 expression in biopsy specimen had significantly lower prostate-specific antigen recurrence-free survival and overall survival than those with no GPX2 expression. These findings suggest that GPX2 is a prognostic marker in CRPC and affects proliferation of prostate cancer under androgen depletion partially through protection against ROS signaling.