Mitochondrial Stress Signals Induce a Drug Tolerant Persister Phenotype in Triple Negative Breast Cancer Cell Models

Abstract

Achieving effective chemotherapy has been a long-standing challenge. Over time cancer cells develop the ability to survive these cytotoxic insults, usually by undergoing genetic mutations leading to drug resistance. In recent years, a distinct pathway involving formation of drug tolerant persister (DTP) cancer cells has been identified. DTP cells utilize novel non-genetic survival mechanisms driven by transcriptional and epigenetic changes without alteration of the cellular genetic makeup. DTP cells have also been shown to modulate innate immune pathways promoting survival instead of cell death. This innate immune activation takes place via pathogen recognition receptors (PRRs) that sense intracellular damaged self-DNA leading to heightened interferon production. Here, we explored the possibility of interferon induction downstream of mitochondrial damage as a novel DTP mechanism. For this, different functional aspects of mitochondria were targeted in human (MDA-MB-231) and mouse (4T1) cell models of triple negative breast cancer including inhibition of ETC complex-1 using IACS-010759. Through transcriptomic analyses, following ETC inhibition, we observed high expression of interferon signalling, which was accompanied by downregulation of 13 genes encoded within mitochondrial DNA (mtDNA). We also directly detected a reduction in mtDNA abundance, with a concomitant loss of mtDNA encoded COX1 protein. IACS-010759 treatment led to an enrichment in fusion-deficient short isoforms of mitochondrial fusion protein OPA1 and transient knockdown of OPA1 also resulted in a reduction in mtDNA encoded genes. Furthermore, CRISPR-Cas9 mediated knockout of OPA1 in HeLa cells significantly reduced mtDNA abundance, confirming that OPA1 is required to maintain mtDNA stability. Taken together, we propose a new mechanism of cancer cell persistence through increased interferon expression during mitochondrial stress. Our findings suggest that mitochondrial stress, such as inhibition of ETC function, can increase fusion-deficient short isoforms of OPA1 disrupting the fusion machinery, which thereby cause release of mitochondrial nucleic acids into the cytoplasm.