Dissociable Rpb4-Rpb7 Subassembly of RNA Polymerase II Binds to Single-strand Nucleic Acid and Mediates a Post-recruitment Step in Transcription Initiation

• Published in: The Journal of biological chemistry Vol. 276; no. 13; pp. 10097 - 10102
• Main Authors: Orlicky, Stephen M., Tran, Phan T., Sayre, Michael H., Edwards, Aled M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 30.03.2001; American Society for Biochemistry and Molecular Biology
• Subjects: Amino Acid Motifs; Amino Acid Sequence; Animals; Baculoviridae - metabolism; Cell Line; DNA, Single-Stranded - metabolism; Dose-Response Relationship, Drug; Escherichia coli - enzymology; Gene Deletion; Insecta; Models, Molecular; Molecular Sequence Data; Mutation; Phenotype; Promoter Regions, Genetic; Protein Binding; Protein Structure, Tertiary; RNA Polymerase II - metabolism; Rpb4 protein; Rpb7 protein; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins - metabolism; Sequence Homology, Amino Acid; Sulfolobus - enzymology; Transcription, Genetic
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M003165200

The Rpb4 and Rpb7 subunits of yeast RNA polymerase II form a heterodimeric complex essential for promoter-directed transcription initiation in a reconstituted system. Results of template competition experiments indicate that the Rpb4-Rpb7 complex is not required for stable recruitment of polymerase to active preinitiation complexes, suggesting that Rpb4-Rpb7 mediates an essential step subsequent to promoter binding. Sequence and structure-based alignments revealed a possible OB-fold single-strand nucleic acid-binding motif in Rpb7. Purified Rpb4-Rpb7 complex exhibited both single-strand DNA- and RNA-binding activities, and a small deletion in the putative OB-fold nucleic acid-binding surface of Rpb7 abolished binding activity without affecting the stability of the Rpb4-Rpb7 complex or its ability to associate with polymerase. The same mutation destroyed the transcription activity of the Rpb4-Rpb7 complex. A separate deletion elsewhere in the OB-fold motif of Rpb7 also blocked transcription but did not affect nucleic acid binding, suggesting that the OB-fold of Rpb7 mediates both DNA-protein and protein-protein interactions required for productive initiation.