Acrolein preferentially damages nucleolus eliciting ribosomal stress and apoptosis in human cancer cells

Abstract

Acrolein (Acr) is a potent cytotoxic and DNA damaging agent which is ubiquitous in the environment and abundant in tobacco smoke. Acr is also an active cytotoxic metabolite of the anti-cancer drugs cyclophosphamide and ifosfamide. The mechanisms via which Acr exerts its anti-cancer activity and cytotoxicity are not clear. In this study, we found that Acr induces cytotoxicity and cell death in human cancer cells with different activities of p53. Acr preferentially binds nucleolar ribosomal DNA (rDNA) to form Acr-deoxyguanosine adducts, and induces oxidative damage to both rDNA and ribosomal RNA (rRNA). Acr triggers ribosomal stress responses, inhibits rRNA synthesis, reduces RNA polymerase I binding to the promoter of rRNA gene, disrupts nucleolar integrity, and impairs ribosome biogenesis and polysome formation. Acr causes an increase in MDM2 levels and phosphorylation of MDM2 in A549 and HeLa cells which are p53 active and p53 inactive, respectively. It enhances the binding of ribosomal protein RPL11 to MDM2 and reduces the binding of p53 and E2F-1 to MDM2 resulting in stabilization/activation of p53 in A549 cells and degradation of E2F-1 in A549 and HeLa cells. We propose that Acr induces ribosomal stress which leads to activation of MDM2 and RPL11-MDM2 binding, consequently, activates p53 and enhances E2F-1 degradation, and that taken together these two processes induce apoptosis and cell death.

Triptolide interrupts rRNA synthesis and induces the RPL23‑MDM2‑p53 pathway to repress lung cancer cells

“…Inhibition of B23 in atypical teratoid or rhabdoid tumors arrests the cell cycle at G1 phase (27), suggesting that reduced expression and abnormal distribution of NCL and B23 induced by TP are implicated in cell cycle arrest, apoptosis and growth inhibition. These results are consistent with previous findings that ribosomal stress caused by acrolein induced nucleolar disintegration and nucleolus-related molecule translocation (16).…”

“…The nucleolus is a pressure sensor that contains various proteins involved in ribosome synthesis (25). B23, NCL, RNA Pol I and UBF are the main sensory proteins located in the nucleoli (16). In the present study, it was found that B23, NCL, RNA Pol I and UBF were located primarily in the nucleolus in control-treated NSCLC cells, whereas in the TP-treated cells, the nucleolar structure disappeared, and B23, NCL, RNA Pol I and UBF were translocated and dispersed throughout the nucleus.…”

“…2A and B. 45S rRNA is the transcriptional product driven by Pol I and UBF, and maturation of the primary transcript leads to the generation of 18S rRNA (16). RT-qPCR analysis was performed to determine the effect of TP on primary rRNA transcript 45S and modified 18S.…”

“…UBF binds to the whole length of the rDNA transcript units and is implicated in the control of the elongation process (15). When cells are subjected to stress, the nucleolus acts as a sensor for cellular stress signals, resulting in the recruitment of ribosomal proteins (RPs) to mouse double minute 2 protein (MDM2), thereby disrupting the interaction between p53 and MDM2 (16). As a result, p53 is no longer degraded by MDM2-mediated ubiquitination, which further induces cell cycle arrest or apoptosis (16).…”

“…When cells are subjected to stress, the nucleolus acts as a sensor for cellular stress signals, resulting in the recruitment of ribosomal proteins (RPs) to mouse double minute 2 protein (MDM2), thereby disrupting the interaction between p53 and MDM2 (16). As a result, p53 is no longer degraded by MDM2-mediated ubiquitination, which further induces cell cycle arrest or apoptosis (16). In addition, TP inhibits the activity of RNA Pol I (17), suggesting that TP may be involved in regulating ribosome biogenesis.…”