Mitochondrial p32-mediated regulation of p-TBK1 affects the radiosensitivity of colorectal cancer.

Purpose: Many colorectal cancer (CRC) patients respond poorly to radiotherapy due to radioresistance. Understanding the molecular mechanisms underlying this resistance is crucial. It was demonstrated that p32, a mitochondrial protein translation regulator, is related to cancer development. However, its specific function and mechanism in CRC, has not yet been investigated. This study aims to explore the role of p32 in CRC and its impact on radiotherapy sensitivity.

Methods: Cell viability was evaluated by MTT and EdU assay. Mitochondrial DNA (mtDNA) leakage was quantified by RT-qPCR. Radiosensitivity was indicated by cellular phosphorylation of H2AX (γH2AX) foci, phosphorylation of ataxia telangiectasia mutated (p-ATM) and phosphorylation of checkpoint kinase 2 (p-CHK2) levels, as well as by mice tumor model subjected to radiotherapy. Moreover, histological and transcriptomic analysis of p32 expression were performed in CRC patients.

Results: In p32-KO cells, we observed reduced cell viability, damaged mitochondria, mtDNA leakage, and increased radiosensitivity. Furthermore, depletion of p32 induced the DNA damage response (DDR) by activating cytoplasmic DNA sensing cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING)-TANK-binding kinase 1 (TBK1), which was reversed by p32/TBK1 double knockout. Depletion of p32 also induced the mitochondrial fragmentation, induced mtDNA leakage through the mitochondrial permeability transition pore (mPTP), effects that could be mitigated by Mdivi-1 or Cyclosporin A (CsA).

Conclusions: Our study demonstrates that inhibiting p32 in CRC enhances radiosensitivity by causing mitochondrial dysfunction, increasing mitochondrial fission, inducing mtDNA leakage and activating the cGAS-STING-TBK1 pathway. These findings provide a potential therapeutic target for overcoming radioresistance in CRC.