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, M(1)dG. The exocyclic ring of M(1)dG opens to the acyclic N(2)-OPdG structure when paired with C but remains closed in single-stranded DNA or when mispaired with T. M(1)dG 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 M(1)dG 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. M(1)dG in the transcribed strand arrested 60-90% POLRMT elongation complexes with greater arrest by the adduct when opposite T. POLRMT was more sensitive to N(2)-OPdG and M(1)dG after initiation at LSP, which suggests promoter-specific differences in the function of POLRMT complexes. A closed-ring analog of M(1)dG, 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 M(1)dG adducts in mitochondrial DNA hinder the transcription of mitochondrial genes.