Restraining Akt1 Phosphorylation Attenuates the Repair of Radiation-Induced DNA Double-Strand Breaks and Reduces the Survival of Irradiated Cancer Cells

Abstract

The survival kinase protein kinase B (Akt) participates in the regulation of essential subcellular processes, e.g., proliferation, growth, survival, and apoptosis, and has a documented role in promoting resistance against genotoxic stress including radiotherapy, presumably by influencing the DNA damage response and DNA double-strand break (DSB) repair. However, its exact role in DSB repair requires further elucidation. We used a genetic approach to explore the consequences of impaired phosphorylation of Akt1 at one or both of its key phosphorylation sites, Threonine 308 (T308) or Serine 473 (S473), on DSB repair and radiosensitivity to killing. Therefore, we overexpressed either the respective single or the double phosphorylation-deficient mutants (Akt1-T308A, Akt1-S473A, or Akt1-T308A/S473A) in TRAMPC1 murine prostate cancer cells (TrC1) and measured the DSB repair kinetics and clonogenic cell survival upon irradiation. Only the expression of the Akt1-T308A/S473A induced a significant delay in the kinetics of DSB repair in irradiated TrC1 as determined by the γH2A.X (H2A histone family, member X) assay and the neutral comet assay, respectively. Moreover, Akt1-T308A/S473A-expressing cells were characterized by increased radiosensitivity compared to Akt1-WT (wild type)-expressing cells in long-term colony formation assays. Our data reveal that Akt1’s activation state is important for the cellular radiation response, presumably by modulating the phosphorylation of effector proteins involved in the regulation of DSB repair.

Regulation of tumor cell – Microenvironment interaction by the autotaxin-lysophosphatidic acid receptor axis

“…These two kinases play key roles in the regulation of cell survival by inhibiting apoptosis. The central role of the Akt kinase–NFkB transcription factor is well documented in mediating resistance to radiation and chemotherapy (Oeck et al, 2017; Szymonowicz et al, 2018). We hypothesize that the upregulated ATX-LPA 2 receptor axis via the IEX-1 scaffold protein (de Laval et al, 2014) and ERK1/2 are important amplifiers of the activity of Akt-NFkB.…”

Inhibition of AKT-Signaling Sensitizes Soft Tissue Sarcomas (STS) and Gastrointestinal Stromal Tumors (GIST) to Doxorubicin via Targeting of Homology-Mediated DNA Repair

“…Importantly, direct interaction between DNA-PK and AKT in cisplatin-resistant but not sensitive cells was revealed by immunoprecipitation [ 25 ]. The regulatory role of DNA-PK in Akt-S473 phosphorylation was also shown for glioblastoma cells in response to IR [ 26 ]. The role of AKT in DNA-PKcs-dependent DNA DSB repair has been extensively studied by Toulany M. with co-authors.…”

Insulin-like growth factor receptor signaling in tumorigenesis and drug resistance: a challenge for cancer therapy

“…Inhibition of IGF-IR impairs the repair of radiation-induced double strand breaks and sensitizes cancer cells to radiotherapy [82]. The promotion of NHEJ by IGF-IR may rely on DNAdependent protein kinase (DNA-PK), whereas a direct interaction between IGF-IR and DNA-PK is absent [93]. Of note, the IGF-IR downstream effectors Akt and mTOR promote the repair of radiation-induced DNA double strand breaks and confer resistance to radiotherapy [93,94].…”

“…The promotion of NHEJ by IGF-IR may rely on DNAdependent protein kinase (DNA-PK), whereas a direct interaction between IGF-IR and DNA-PK is absent [93]. Of note, the IGF-IR downstream effectors Akt and mTOR promote the repair of radiation-induced DNA double strand breaks and confer resistance to radiotherapy [93,94]. Inhibition of residual homologous recombination further enhances the sensitivity to IGF-IR inhibitors [95].…”