title: SpoIIAA governs the release of the cell-type specific transcription factor sigma F from its anti-sigma factor SpoIIAB.
abstract: The Bacillus subtilis transcription factor sigma F is a cell-type specific regulatory protein whose activity is governed by SpoIIAB and SpoIIAA and the nucleotides ATP and ADP. SpoIIAB is an anti-sigma factor that binds to sigma F in a manner that is stimulated by ATP, thereby trapping sigma F in an inactive complex. Alternatively, SpoIIAB binds to SpoIIAA in a manner that is stimulated by ADP to form a SpoIIAB.SpoIIAA complex in which SpoIIAB is sequestered from sigma F. SpoIIAB is also a protein kinase that uses ATP to phosphorylate, and thereby inactivate, SpoIIAA. Thus, ATP inhibits sigma F activity both by promoting formation of the SpoIIAB.sigma F complex and by phosphorylation of SpoIIAA. In extension of previous results, we use affinity chromatography to show that SpoIIAB is capable of forming long-lived complexes with sigma F and SpoIIAA and that the formation of these complexes is dependent on ATP and ADP, respectively. Using a DNA template lacking adenosine residues on the non-transcribed strand, we demonstrate that ATP is required for SpoIIAB-mediated inhibition of sigma F-directed RNA synthesis and that this inhibition is prevented by SpoIIAA in a manner that is stimulated by ADP. We show that ADP acts by protecting SpoIIAA from phosphorylation by SpoIIAB and that a mutant protein bearing an amino acid substitution at the site of phosphorylation in SpoIIAA is capable of preventing the inhibition of sigma F in a manner that does not depend on ADP. A principal finding from the investigation is that SpoIIAA restores activity to sigma F that had previously been inhibited by SpoIIAB. This is demonstrated both by the capacity of SpoIIAA to reverse SpoIIAB-mediated inhibition of sigma F-directed RNA synthesis and by its capacity to interact with and disrupt the SpoIIAB. sigma F complex. The results are consistent with a model in which sigma F is controlled by the cellular concentration of unphosphorylated SpoIIAA.
authors.affiliation: Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, USA.
authors.forename: L
authors.initials: L
authors.lastname: Duncan
authors.forename: S
authors.initials: S
authors.lastname: Alper
authors.forename: R
authors.initials: R
authors.lastname: Losick
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology; D001412:Bacillus subtilis / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D008957:Models, Genetic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D010766:Phosphorylation; D011485:Protein Binding; D011487:Protein Conformation; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; C064971:FliA protein, Bacteria; D012808:Sigma Factor; D014157:Transcription Factors; C092468:spoIIR protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate