title: FliT selectively enhances proteolysis of FlhC subunit in FlhD4C2 complex by an ATP-dependent protease, ClpXP.
abstract: We previously reported that the ClpXP ATP-dependent protease specifically recognizes and degrades the flagellar master transcriptional activator complex, FlhD4C2, to negatively control flagellar biogenesis. The flagellum-related protein, FliT, is also a negative regulator of flagellar regulon by inhibiting the binding of FlhD4C2 to the promoter DNA. We have found a novel pathway of FliT inhibition of FlhD4C2 activity connected to ClpXP proteolysis. An in vitro degradation assay using purified proteins shows that FliT selectively increases ClpXP proteolysis of the FlhC subunit in the FlhD4C2 complex. FliT behaves specifically to ClpXP-dependent proteolysis of FlhC. An in vitro interaction assay detects the ternary complex of FliT-FlhD4C2-ClpX. FliT promotes the affinity of ClpX against FlhD4C2 complex, whereas FliT does not directly interact with ClpX. Thus, FliT interacts with the FlhC in FlhD4C2 complex and increases the presentation of the FlhC recognition region to ClpX. The DNA-bound form of FlhD4C2 complex is resistant to ClpXP proteolysis. We suggest that the role of FliT in negatively controlling the flagellar gene expression involves increasing free molecules of FlhD4C2 sensitive to ClpXP proteolysis by inhibiting the binding to the promoter DNA as well as enhancing the selective proteolysis of FlhC subunit by ClpXP.
authors.affiliation: From the Department of Microbiology and Molecular Genetics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, 260-8675 Japan and.
authors.forename: Yoshiharu
authors.initials: Y
authors.lastname: Sato
authors.affiliation: From the Department of Microbiology and Molecular Genetics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, 260-8675 Japan and.
authors.forename: Akiko
authors.initials: A
authors.lastname: Takaya
authors.affiliation: the Department of Biology, University of Utah, Salt Lake City, Utah 84112.
authors.forename: Chakib
authors.initials: C
authors.lastname: Mouslim
authors.affiliation: the Department of Biology, University of Utah, Salt Lake City, Utah 84112 kelly.hughes@utah.edu.
authors.forename: Kelly T
authors.initials: KT
authors.lastname: Hughes
authors.affiliation: From the Department of Microbiology and Molecular Genetics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba, 260-8675 Japan and tomoko-y@faculty.chiba-u.jp.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Yamamoto
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D049071:Endopeptidase Clp / Q000235:genetics / Q000378:metabolism*; D005407:Flagella / Q000378:metabolism; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D021122:Protein Subunits / Q000235:genetics / Q000378:metabolism; D059748:Proteolysis; D018085:Regulon; D012486:Salmonella typhimurium / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004269:DNA, Bacterial; D018832:Molecular Chaperones; D021122:Protein Subunits; D015534:Trans-Activators; C076735:fliT protein, Bacteria; D049071:Endopeptidase Clp
keywords: ATPases Associated with Diverse Cellular Activities (AAA); Chaperone; Gene Regulation; Protein Degradation; Proteolysis