Dimer formation and transcription activation in the sporulation response regulator Spo0A.
The response regulator Spo0A is the master control element in the initiation of sporulation in Bacillus subtilis. Like many other multi-domain response regulators, the latent activity of the effector, C-terminal domain is stimulated by phosphorylation on a conserved aspartic acid residue in the regulatory, N-terminal domain. If a threshold concentration of phosphorylated Spo0A is achieved, the transcription of genes required for sporulation is activated, whereas the genes encoding stationary phase sentinels are repressed, and sporulation proceeds. Despite detailed genetic, biochemical and structural characterisation, it is not understood how the phosphorylation signal in the receiver domain is transduced into DNA binding and transcription activation in the distal effector domain. An obstacle to our understanding of Spo0A function is the uncertainty concerning changes in quaternary structure that accompany phosphorylation. Here we have revisited this question and shown unequivocally that Spo0A forms dimers upon phosphorylation and that the subunit interactions in the dimer are mediated principally by the receiver domain. Purified dimers of two mutants of Spo0A, in which the phosphorylatable aspartic acid residue has been substituted, activate transcription from the spoIIG promoter in vitro, whereas monomers do not. This suggests that dimers represent the activated form of Spo0A.
Lewis
Richard J
RJ
Structural Biology Laboratory, Department of Chemistry, University of York, YO10 5DD, UK.
Scott
David J
DJ
Brannigan
James A
JA
Ladds
Joanne C
JC
Cervin
Marguerite A
MA
Spiegelman
George B
GB
Hoggett
James G
JG
Barák
Imrich
I
Wilkinson
Anthony J
AJ
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics* / Q000502:physiology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002850:Chromatography, Gel; D018360:Crystallography, X-Ray; D004247:DNA / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial / Q000235:genetics; D005809:Genes, Regulator / Q000235:genetics; D008954:Models, Biological; D008958:Models, Molecular; D008970:Molecular Weight; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits; D012808:Sigma Factor; D013171:Spores, Bacterial / Q000235:genetics*; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014461:Ultracentrifugation
D001426:Bacterial Proteins; D021122:Protein Subunits; D012808:Sigma Factor; C080950:Spo0A protein, Bacillus subtilis; D014157:Transcription Factors; C092468:spoIIR protein, Bacillus subtilis; C030069:spore-specific proteins, Bacillus; D004247:DNA; D010744:Phosphoric Monoester Hydrolases