Arrest of human mitochondrial RNA polymerase transcription by the biological aldehyde adduct of DNA, M₁dG

The biological aldehydes, malondialdehyde and base propenal, react with DNA to form a prevalent guanine adduct, M₁dG. The exocyclic ring of M₁dG opens to the acyclic N²-OPdG structure when paired with C but remains closed in single-stranded DNA or when mispaired with T. M₁dG is a target of nucleotide excision repair (NER); however, NER is absent in mitochondria. An in vitro transcription system with purified human mitochondrial RNA polymerase (POLRMT) and transcription factors, mtTFA and mtTFB2, was used to determine the effect of M₁dG on POLRMT elongation. DNA templates contained a single adduct opposite either C or T downstream of either the light-strand (LSP) or heavy-strand (HSP1) promoter for POLRMT. M₁dG in the transcribed strand arrested 60-90% POLRMT elongation complexes with greater arrest by the adduct when opposite T. POLRMT was more sensitive to N²-OPdG and M₁dG after initiation at LSP, which suggests promoter-specific differences in the function of POLRMT complexes. A closed-ring analog of M₁dG, PdG, blocked ≥95% of transcripts originating from either promoter regardless of base pairing, and the transcripts remained associated with POLRMT complexes after stalling at the adduct. This work suggests that persistent M₁dG adducts in mitochondrial DNA hinder the transcription of mitochondrial genes.