An ATP-dependent partner switch links flagellar C-ring assembly with gene expression

Abstract

Bacterial flagella differ in their number and spatial arrangement. In many species, the MinD-type ATPase FlhG (also YlxH/FleN) is central to the numerical control of bacterial flagella, and its deletion in polarly flagellated bacteria typically leads to hyperflagellation. The molecular mechanism underlying this numerical control, however, remains enigmatic. Using the model species Shewanella putrefaciens, we show that FlhG links assembly of the flagellar C ring with the action of the master transcriptional regulator FlrA (named FleQ in other species). While FlrA and the flagellar C-ring protein FliM have an overlapping binding site on FlhG, their binding depends on the ATP-dependent dimerization state of FlhG. FliM interacts with FlhG independent of nucleotide binding, while FlrA exclusively interacts with the ATP-dependent FlhG dimer and stimulates FlhG ATPase activity. Our in vivo analysis of FlhG partner switching between FliM and FlrA reveals its mechanism in the numerical restriction of flagella, in which the transcriptional activity of FlrA is down-regulated through a negative feedback loop. Our study demonstrates another level of regulatory complexity underlying the spationumerical regulation of flagellar biogenesis and implies that flagellar assembly transcriptionally regulates the production of more initial building blocks.

Dynamic Hybrid Flagellar Motors—Fuel Switch and More

“…In agreement with cryo–electron tomography studies, the data indicate that, in the S. oneidensis motor, eleven stators can be synchronously engaged ( Kaplan et al, 2019 ; Blagotinsek et al, 2020 ). In the presence of Na + , eight of the 11 positions are occupied by the Na + -dependent PomAB stator.…”

Interdependent Polar Localization of FlhF and FlhG and Their Importance for Flagellum Formation of Vibrio parahaemolyticus

“…It would be interesting to study whether a similar regulatory mechanism exists in V. parahaemolyticus . We find it noteworthy that FlhG directly interacts with a transcriptional regulator ( Blagotinsek et al, 2020 ) and we speculate that perhaps its localization in non-polar foci, which we observe in the absence of HubP, could be related to its function in transcriptional regulation and possibly reflect an interaction with transcriptional regulators on the chromosome – hereby giving rise to the distinct focus localization sites that are particularly enhanced in the absence of HubP in V. parahaemolyticus . In this way, we would like to hypothesize that the localization of FlhG to the cell pole might not only reflect a function in regulating FlhF activity, but possibly to sequester it spatially to prevent its action on transcriptional regulation in the cytoplasm as a specific cell cycle check point.…”

“…In V. cholerae , both FlhF and FlhG are known transcriptional regulators of flagellar genes ( Correa et al, 2005 ). Indeed, very recently it was shown that FlhG plays a very direct role in regulating the expression of flagellum genes in S. putrefaciens by connecting the initial phases of flagellum formation with the activity of the transcriptional regulator FlrA ( Blagotinsek et al, 2020 ), which in V. parahaemolyticus is referred to as FlaK. It would be interesting to study whether a similar regulatory mechanism exists in V. parahaemolyticus .…”

“…It is essential to understand the mechanisms required for dissemination of bacteria in the environment and for many bacteria, the primary means of motion is flagella-mediated swimming motility. Correct swimming behavior heavily depends on the production of the correct number and proper placement of the flagella within the cell ( Schuhmacher et al, 2015b ; Blagotinsek et al, 2020 ; Kojima et al, 2020 ).…”

Dynamics of Bacterial Signal Recognition Particle at a Single Molecule Level

“…In order to analyze if changes seen for translation inhibition were specific for Ffh, FtsY, and ribosomes, we visualized the dynamics of a protein that is unrelated to translation. We chose the enhancer binding protein FlrA, which plays an important role in gene regulation for establishment of the single polar flagellum in Shewanella (Blagotinsek et al, 2020). FlrA is a hexameric AAA + protein that influences RNA polymerase activity (Gao et al, 2020).…”