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MRE11 promotes AKT phosphorylation in direct response to DNA double-strand breaks
Michael Fraser1, Shane M Harding, Helen Zhao
1Campbell Family Cancer Research Institute/Ontario Cancer Institute, Toronto, Ontario, Canada.
Abstract:
AKT is hyper-activated in many human cancers and promotes proliferation and cancer cell survival in response to DNA damaging agents. Ionizing radiation (IR) produces DNA double strand breaks (DSB) and activates AKT, however a direct mechanism linking intra-nuclear DSB and AKT signaling is lacking. Here we demonstrate that AKT is phosphorylated following IR in benign and malignant cells and, using colony-forming assays and in vitro rejoining assays, show that AKT promotes non-homologous end joining-mediated DSB repair and cell survival following IR. Further studies revealed that pAKT-S473, but not pAKT-T308 or total AKT, accumulates in the vicinity of IR-induced DSB and co-localizes with γH2AX and ATM-pSer1981. Based on whole-cell IR, nuclear UV microbeam, and endonuclease-induced DSB studies, we observed that pAKT-S473 is up-regulated by a DSB-induced signaling cascade, and this is dependent on the DSB sensor protein, MRE11. MRE11-dependent pAKT-S473 did not require the MRE11 endonuclease domain. The histone ubiquitin ligase RNF168 is also required for DSB-induced pAKT-S473, and DSB-induced pAKT-S473 is independent of DNA-PKcs, PI3K, and ATR. These data demonstrate that DSB activate a signaling cascade that directly promotes a PI3K-independent pathway of AKT phosphorylation that is dependent on MRE11-ATM-RNF168 signaling. Thus, these data directly link the presence of DNA breaks to AKT-mediated cell survival and support AKT as a target for cancer therapy.
Insights
DNA double-strand breaks (DSB) activate AKT signaling, promoting cancer cell survival and DNA repair. This study reveals a novel MRE11-ATM-RNF168 pathway linking DSB to AKT phosphorylation, independent of PI3K, highlighting AKT as a potential cancer therapy target.
Area of Science:
- Molecular Biology
- Cancer Research
- Cell Signaling
Background:
- Hyper-activated AKT signaling is common in human cancers, promoting cell proliferation and survival, particularly in response to DNA damage.
- Ionizing radiation (IR) induces DNA double-strand breaks (DSB) and activates AKT, but the direct molecular link between nuclear DSB and AKT signaling remains unclear.
Purpose of the Study:
- To elucidate the direct mechanism linking intra-nuclear DNA double-strand breaks (DSB) to AKT signaling activation.
- To investigate the role of AKT in DNA double-strand break repair and cell survival following ionizing radiation (IR).
Main Methods:
- Utilized colony-forming and in vitro rejoining assays to assess AKT's role in DSB repair and cell survival post-IR.
- Employed techniques including whole-cell IR, nuclear UV microbeam, and endonuclease-induced DSB studies to analyze pAKT-S473 localization and regulation.
- Investigated the involvement of key proteins such as MRE11, ATM, RNF168, DNA-PKcs, PI3K, and ATR in the DSB-induced AKT phosphorylation cascade.
Main Results:
- Demonstrated that AKT is phosphorylated following IR in both benign and malignant cells, promoting non-homologous end joining (NHEJ)-mediated DSB repair and cell survival.
- Showed that phosphorylated AKT at serine 473 (pAKT-S473) accumulates near IR-induced DSB sites and co-localizes with DSB markers like γH2AX and ATM-pSer1981.
- Identified a novel DSB-induced signaling cascade dependent on MRE11, ATM, and RNF168 that leads to PI3K-independent AKT phosphorylation at S473.
Conclusions:
- Established a direct link between the presence of DNA double-strand breaks and AKT-mediated cell survival.
- The findings reveal a novel PI3K-independent pathway for AKT phosphorylation triggered by DSB through MRE11-ATM-RNF168 signaling.
- Support targeting AKT as a viable strategy for cancer therapy, given its crucial role in DSB repair and cell survival in response to DNA damage.
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