Autophagy-Mediated Clearance of free Genomic DNA in the Cytoplasm Protects the Growth and Survival of Cancer Cells

Abstract Background: The cGAS (GMP-AMP synthase)-triggered senescence-associated secretory phenotype (SASP) in promotion of cancer progression has been extensively documented. However, the role of cGAS-mediated DNA autophagy is little evaluated in cancer cells.Methods: Immunofluorescence, senescence associated-β-galactosidase staining (SA-β-gal) and Western blot were performed to detect gene expression, distribution and phenotypes. PCR, IP-PCR, FISH, BrdU, Comet assay, coimmunoprecipitation, sucrose density gradient centrifugation were carried out to detect possible mechanisms. Trypan blue exclusion, Live/dead staining and MTS assay were to measure the cell viability. All analyses were performed using GraphPad Prism 8. Relationships were analyzed using t-tests. A P-value of less than 0.05 was considered significant. All statistical tests and P values were 2-sided, and the level of significance was set at <0.05 (*), <0.01 (**), <0.001 (***), or<0.0001 (****); ns indicates no significance.Results: Active DNA autophagy but not SASP activity could be detected in the BT-549 breast cancer cells with high micronucleus (MN). The selective autophagy of free genomic DNA in the cytoplasm is mediated by cGAS and usually coordinated with SQSTM1-mediated autophagy of ubiquitinated histones. Cytoplasmic DNA together with nuclear proteins derive from DNA replication-induced nuclear damage and MN collapse. The inhibition of DNA autophagy through either chemical inhibitors or genomic silencing of cGAS or SQSTM1 suppresses the growth and survival of cancer cells, while enhanced DNA damage increases the sensitivity to these inhibitors. Human cancer cells with either high DNA autophagy or enhancement of DNA damage are sensitive to inhibition of DNA autophagy.Conclusions: Our investigation revealed DNA autophagy in breast cancer cells with high MN formation. Autophagy of genomic DNA in the cytosol could be mediated by cGAS but is usually coordinated with other autophagic mediators. The selective autophagy mediated clearance of free genomic DNA protects of growth and survival cancer cells, and autophagic inhibition could be a potential therapeutic approach for cancer cells with high DNA autophagic activity.