Differential modulation of cancer-related genes by mitochondrial DNA haplogroups and the STING DNA sensing system

Abstract

Activation of the Simulator of Interferon Genes (STING) system by mitochondrial (mt) DNA can upregulate type 1 interferon genes and enhance immune responses to combat bacterial and viral infections. In cancers, the tumor-derived DNA activates STING leading to upregulation of IFN-beta and induction of antitumor T cells. The entire mtDNA from the cell lines was sequenced using next-generation sequencing (NGS) technology with independent sequencing of both strands in both directions, allowing identification of low-frequency heteroplasmy SNPs. There were 15 heteroplasmy SNPs showing a range from 3.4% to 40.5% occurrence in the K cybrid cell lines. Three H haplogroup cybrids possessed SNP heteroplasmy that ranged from 4.39% to 30.7%. The present study used qRT-PCR to determine if cybrids of H and K haplogroups differentially regulate expression levels of five cancer genes (BRAC1, ALK, PD1, EGFR, and HER2) and seven STING subunits genes (CGAS, TBK1, IRF3, IκBa, NFκB, TRAF2, and TNFRSF19). Some cybrids underwent siRNA knockdown of STING followed by qRT-PCR in order to determine the impact of STING on gene expression. Rho0 (lacking mtDNA) ARPE-19 cells were used to determine if mtDNA is required for the expression of the cancer genes studied. Our results showed that (a) K cybrids have lower expression levels for BRAC1, ALK, PD1, EGFR, IRF3, and TNFRSF19 genes but increased transcription for IκBa and NFκB compared to H cybrids; (b) STING KD decreases expression of EGFR in both H and K cybrids, and (c) PD1 expression is negligible in Rho0 cells. Our findings suggest that the STING DNA sensing pathway may be a previously unrecognized pathway to target modulation of cancer-related genes and the PD1 expression requires the presence of mtDNA.