A DNA-tethered cleavage probe reveals the path for promoter DNA in the yeast preinitiation complex

• Published in: Nature structural & molecular biology Vol. 13; no. 7; pp. 603 - 610
• Main Authors: Hahn, Steven, Miller, Gail
• Format: Journal Article
• Language: English
• Published: United States Nature Publishing Group 01.07.2006
• Subjects: Base Sequence; Binding sites; Crystal structure; Deoxyribonucleic acid; DNA; DNA Helicases - chemistry; DNA Helicases - genetics; DNA Helicases - metabolism; DNA Polymerase II - chemistry; DNA Polymerase II - genetics; DNA probes; DNA, Fungal - chemistry; DNA, Fungal - genetics; Genetic transcription; Genomics; Hydroxyl Radical; Models, Molecular; Nucleic Acid Conformation; Physiological aspects; Promoter Regions, Genetic; Promoters (Genetics); Ribonucleic acid; RNA; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - chemistry; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Transcription Factor TFIIH - chemistry; Transcription Factor TFIIH - genetics; Transcription, Genetic; Yeast; Yeasts; United States
• ISSN: 1545-9993; 1545-9985
• DOI: 10.1038/nsmb1117

To directly map the position of promoter DNA within the RNA polymerase II (Pol II) transcription preinitiation complex (PIC), FeBABE was tethered to specific sites within the HIS4 promoter and used to map exposed surfaces of Pol II and the general transcription factors in proximity to DNA. Our results distinguish between previously proposed models for PIC structure and demonstrate that downstream promoter DNA is positioned over the central cleft of Pol II, with DNA upstream of TATA extending toward the Pol II subunit Rpb3. Also mapped were segments of TFIIB, TFIIE, TFIIF and TFIIH in proximity to promoter DNA. DNA downstream of the transcription bubble maps to a path between the two helicase subdomains of the TFIIH subunit Rad25 (also called XPB). Together, our results show how the general factors and Pol II converge on promoter DNA within the PIC.