The phosphorylation state of the DegU response regulator acts as a molecular switch allowing either degradative enzyme synthesis or expression of genetic competence in Bacillus subtilis.
Two classes of mutations were identified in the degS and degU regulatory genes of Bacillus subtilis, leading either to deficiency of degradative enzyme synthesis (degS or degU mutations) or to a pleiotropic phenotype which includes overproduction of degradative enzymes and the loss of genetic competence (degS(Hy) or degU(Hy) mutations). We have shown previously that the DegS protein kinase and the DegU response regulator form a signal transduction system in B. subtilis. We now demonstrate that the DegS protein kinase also acts as a DegU phosphatase. We present evidence that the DegU response regulator has two active conformations: a phosphorylated form which is necessary for degradative enzyme synthesis and a nonphosphorylated form required for expression of genetic competence. The degU146-encoded response regulator, allowing expression of genetic competence, has been purified and seems to be modified within the putative phosphorylation site (D56----N) since it is no longer phosphorylated by DegS. Both the degU146 mutation as well as the degS220 mutation, which essentially abolishes DegS protein kinase activity, lead to deficiency of degradative enzyme synthesis, indicating the requirement of phosphorylated DegU for the expression of this phenotype. We also purified the degU32(Hy)-encoded protein and showed that this response regulator is phosphorylated by the DegS protein kinase in vitro. In addition, the phosphorylated form of the degU32(Hy)-encoded protein presented a strongly increased stability as compared with the wild type DegU protein, thus leading to hyperproduction of degradative enzymes in vivo.
Dahl
M K
MK
Unité de Biochimie Microbienne, Centre National de la Recherche Scientifique URA 1300, Institut Pasteur, Paris, France.
Msadek
T
T
Kunst
F
F
Rapoport
G
G
D000483:Alleles; D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D005798:Genes, Bacterial; D005809:Genes, Regulator; D007700:Kinetics; D008954:Models, Biological; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D010957:Plasmids; D011494:Protein Kinases / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015183:Restriction Mapping; D015398:Signal Transduction
D001426:Bacterial Proteins; D004269:DNA, Bacterial; C066496:DegS protein, Bacteria; C066497:DegU protein, Bacteria; D011994:Recombinant Proteins; D011494:Protein Kinases; D010749:Phosphoprotein Phosphatases