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Linking structural assembly to gene expression: a novel mechanism for regulating the activity of a σ 54 transcription factor
Author(s) -
Dutton Rachel J.,
Xu Zhaohui,
Gober James W.
Publication year - 2005
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2005.04857.x
Subject(s) - caulobacter crescentus , biology , repressor , enhancer , transcription factor , activator (genetics) , mutant , transcription (linguistics) , microbiology and biotechnology , general transcription factor , immunoprecipitation , regulation of gene expression , trans acting , gene , gene expression , promoter , genetics , linguistics , philosophy , bacterial protein
Summary In Caulobacter crescentus , the temporal and spatial expression of late flagellar genes is regulated by the σ 54 transcriptional activator, FlbD. Genetic experiments have indicated that the trans ‐acting factor FliX regulates FlbD in response to the progression of flagellar assembly, repressing FlbD activity until an early flagellar basal body structure is assembled. Following assembly of this structure, FliX is thought to function as an activator of FlbD. Here we have investigated the mechanism of FliX‐mediated regulation of FlbD activity. In vitro transcription experiments showed that purified FliX could function as a repressor of FlbD‐activated transcription. Transcription activated by a gain‐of‐function mutant of FlbD (FlbD‐1204) that is active in vivo in the absence of an early flagellar structure, was resistant to the repressive effects of FliX. DNA binding studies showed that FliX inhibited the interaction of wild‐type FlbD with enhancer DNA but did not effect FlbD‐catalysed ATPase activity. DNA binding activity of FlbD‐1204 was relatively unaffected by FliX indicating that this mutant protein bypasses the transcriptional requirement for early flagellar assembly by escaping FliX‐mediated negative regulation. Gel filtration and co‐immunoprecipitation experiments indicated that FliX formed a stable complex with FlbD. These experiments demonstrate that regulation of FlbD activity is unusual among the well‐studied σ 54 transcriptional activators, apparently combining a two‐component receiver domain with additional control imposed via interaction with a partner protein, FliX.