DNA binding preferences of S. cerevisiae RNA polymerase I Core Factor reveal a preference for the GC-minor groove and a conserved binding mechanism

• Published in: Biochimica et biophysica acta. Gene regulatory mechanisms 2019-09, Vol.1862 (9), p.194408-194408, Article 194408
• Main Authors: Jackobel, Ashleigh J., Zeberl, Brian J., Glover, Danea M., Fakhouri, Aula M., Knutson, Bruce A.
• Format: Article
• Language: English
• Subjects: Benzothiazoles - pharmacology; Conserved Sequence - genetics; Core Factor; CX-5461; DNA binding; DNA, Ribosomal - genetics; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; Electrophoretic Mobility Shift Assay; EMSA; Humans; Minor groove; Naphthyridines - pharmacology; Nucleic Acid Conformation - drug effects; Promoter Regions, Genetic - genetics; RNA polymerase I; RNA Polymerase I - chemistry; RNA Polymerase I - genetics; Saccharomyces cerevisiae - genetics; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription, Genetic
• ISSN: 1874-9399; 1876-4320
• DOI: 10.1016/j.bbagrm.2019.194408

In Saccharomyces cerevisiae, Core Factor (CF) is a key evolutionarily conserved transcription initiation factor that helps recruit RNA polymerase I (Pol I) to the ribosomal DNA (rDNA) promoter. Upregulated Pol I transcription has been linked to many cancers, and targeting Pol I is an attractive and emerging anti-cancer strategy. Using yeast as a model system, we characterized how CF binds to the Pol I promoter by electrophoretic mobility shift assays (EMSA). Synthetic DNA competitors along with anti-tumor drugs and nucleic acid stains that act as DNA groove blockers were used to discover the binding preference of yeast CF. Our results show that CF employs a unique binding mechanism where it prefers the GC-rich minor groove within the rDNA promoter. In addition, we show that yeast CF is able to bind to the human rDNA promoter sequence that is divergent in DNA sequence and demonstrate CF sensitivity to the human specific Pol I inhibitor, CX-5461. Finally, we show that the human Core Promoter Element (CPE) can functionally replace the yeast Core Element (CE) in vivo when aligned by conserved DNA structural features rather than DNA sequence. Together, these findings suggest that the yeast CF and the human ortholog Selectivity Factor 1 (SL1) use an evolutionarily conserved, structure-based mechanism to target DNA. Their shared mechanism may offer a new avenue in using yeast to explore current and future Pol I anti-cancer compounds. •Yeast CF binds preferentially to the GC-minor groove.•A minimal 12 base pair sequence of the yeast CE is required for CF DNA binding.•Yeast CF can bind the human rDNA promoter in a CPE-dependent manner.•CX-5461 inhibits yeast CF from binding to yeast and human rDNA promoters.•The human CPE can functionally replace the yeast CE in vivo.