UV irradiation alters TFAM binding to mitochondrial DNA

Mitochondria lack nucleotide excision repair; however, mitochondrial DNA (mtDNA) is resistant to mutation accumulation following DNA damage. These observations suggest additional damage sensing or protection mechanisms. Transcription Factor A, Mitochondrial (TFAM) compacts mtDNA into nucleoids and binds differentially to certain forms of DNA damage. As such, TFAM has emerged as a candidate for protecting mtDNA or sensing damage. To examine the possibilities that TFAM might protect DNA from damage or act as a damage sensing protein for irreparable forms of mtDNA damage, we used live-cell imaging, cell-based assays, atomic force microscopy (AFM), and high-throughput protein-DNA binding assays to characterize the binding properties of TFAM to ultraviolet-C (UVC) irradiated DNA and the cellular consequences of UVC irradiation. Our cell data show increased TFAM mRNA after exposure and suggest an increase in mtDNA degradation and turnover without a loss in mitochondrial membrane potential that might trigger mitophagy. Our protein-DNA binding assays indicate a reduction in sequence specificity of TFAM following UVC irradiation and a redistribution of TFAM binding throughout the mitochondrial genome. Our AFM data show increased compaction of DNA by TFAM in the presence of damage. Despite the TFAM-mediated compaction of mtDNA, we do not observe any protective effect of increased compaction on DNA damage formation in cells or in vitro. Taken together, these studies indicate that UVC-induced DNA damage alters TFAM binding and promotes compaction by TFAM, suggesting that TFAM may act as a damage sensing protein, sequestering damaged genomes to prevent mutagenesis by facilitating removal or suppression of replication.