title: New insights into transcription fidelity: thermal stability of non-canonical structures in template DNA regulates transcriptional arrest, pause, and slippage.
abstract: The thermal stability and topology of non-canonical structures of G-quadruplexes and hairpins in template DNA were investigated, and the effect of non-canonical structures on transcription fidelity was evaluated quantitatively. We designed ten template DNAs: A linear sequence that does not have significant higher-order structure, three sequences that form hairpin structures, and six sequences that form G-quadruplex structures with different stabilities. Templates with non-canonical structures induced the production of an arrested, a slipped, and a full-length transcript, whereas the linear sequence produced only a full-length transcript. The efficiency of production for run-off transcripts (full-length and slipped transcripts) from templates that formed the non-canonical structures was lower than that from the linear. G-quadruplex structures were more effective inhibitors of full-length product formation than were hairpin structure even when the stability of the G-quadruplex in an aqueous solution was the same as that of the hairpin. We considered that intra-polymerase conditions may differentially affect the stability of non-canonical structures. The values of transcription efficiencies of run-off or arrest transcripts were correlated with stabilities of non-canonical structures in the intra-polymerase condition mimicked by 20 wt% polyethylene glycol (PEG). Transcriptional arrest was induced when the stability of the G-quadruplex structure (-ΔG°37) in the presence of 20 wt% PEG was more than 8.2 kcal mol(-1). Thus, values of stability in the presence of 20 wt% PEG are an important indicator of transcription perturbation. Our results further our understanding of the impact of template structure on the transcription process and may guide logical design of transcription-regulating drugs.
authors.affiliation: Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan.
authors.forename: Hisae
authors.initials: H
authors.lastname: Tateishi-Karimata
authors.affiliation: Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan.
authors.forename: Noburu
authors.initials: N
authors.lastname: Isono
authors.affiliation: Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, Kobe, Japan; Faculty of Frontiers of Innovative Research in Science and Technology (FIRST), Konan University, Kobe, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Sugimoto
mesh_terms: D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D004247:DNA / Q000737:chemistry*; D004591:Electrophoresis, Polyacrylamide Gel; D005453:Fluorescence; D006358:Hot Temperature; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011524:Protoporphyrins / Q000378:metabolism; D013698:Templates, Genetic; D013816:Thermodynamics; D014158:Transcription, Genetic / Q000235:genetics / Q000502:physiology*
chemical_list: D011524:Protoporphyrins; D004247:DNA; C028025:protoporphyrin IX