Gap Junction Intercellular Communication Positively Regulates Cisplatin Toxicity by Inducing DNA Damage through Bystander Signaling

The radiation-induced bystander effect (RIBE) can increase cellular toxicity in a gap junction dependent manner in unirradiated bystander cells. Recent reports have suggested that cisplatin toxicity can also be mediated by functional gap junction intercellular communication (GJIC). In this study using lung and ovarian cancer cell lines, we showed that cisplatin cytotoxicity is mediated by cellular density. This effect is ablated when GJA1 or Connexin 43 (Cx43) is targeted, a gap junction gene and protein, respectively, leading to cisplatin resistance but only at high or gap junction forming density. We also observed that the cisplatin-mediated bystander effect was elicited as DNA Double Strand Breaks (DSBs) with positive H2AX Ser139 phosphorylation (γH2AX) formation, an indicator of DNA DSBs. These DSBs are not observed when gap junction formation is prevented. We next showed that cisplatin is not the "death" signal traversing the gap junctions by utilizing the cisplatin-GG intrastrand adduct specific antibody. Finally, we also showed that cells deficient in the structure-specific DNA endonuclease ERCC1-ERCC4 (ERCC1-XPF), an important mediator of cisplatin resistance, further sensitized when treated with cisplatin in the presence of gap junction forming density. Taken together, these results demonstrate the positive effect of GJIC on increasing cisplatin cytotoxicity.