The Bacillus subtilis Response Regulator Spo0A Stimulates σA-Dependent Transcription Prior to the Major Energetic Barrier

• Published in: The Journal of biological chemistry 2004-04, Vol.279 (17), p.17397-17403
• Main Authors: Seredick, Steve D., Spiegelman, George B.
• Format: Article
• Language: English
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M311190200

At the spoIIG promoter phosphorylated Spo0A (Spo0A∼P) binds 0A boxes overlapping the -35 element, interacting with RNA polymerase to facilitate open complex formation. We have compared in vitro transcription from a series of heteroduplex templates containing denatured regions within the promoters. Transcription from heteroduplex templates with 12, 8, or 6 base pairs denatured was independent of Spo0A∼P, but heteroduplexes with 4 or 2 base pairs denatured required Spo0A∼P for maximal levels of transcription. Investigation of the thermal dependence of transcription suggested that strand separation was the primary thermodynamic barrier to transcription initiation but indicated that Spo0A∼P does not reduce this energetic barrier. Kinetic assays revealed that Spo0A∼P stimulated both the rate of formation of initiated complexes as well as increasing the number of complexes capable of initiating transcription. These results imply that Spo0A∼P stimulates transcription at least in part by stabilizing the RNA polymerase-spoIIG complex until contacts between RNA polymerase and the -10 element induce strand separation.