Spatial, temporal and numerical regulation of polar flagella assembly in Pseudomonas putida

Abstract

The Gram-negative bacterium Pseudomonas putida bears a tuft of flagella at a single cell pole. New flagella must be assembled de novo every cell cycle to secure motility of both daughter cells. Here we show that the coordinated action of FimV, FlhF and FleN sets the location, timing and number of flagella assembled. The polar landmark proteins FimV and FlhF are independently targeted to the nascent new pole during or shortly after cell division, but FimV stabilizes FlhF association with the cell poles. FlhF determines the polar position of the flagella by targeting early flagellar components to the cell pole and preventing their nucleation at non-polar sites. FlhF also promotes efficient flagellar assembly and indirectly stimulates Class III flagellar promoter activation by promoting secretion of the anti-FliA anti-{sigma} factor FlgM. The MinD-like ATPase FleN partitions between the cell poles and the cytoplasm. Cytoplasmic FleN regulates flagellar number by preventing excessive accumulation of FlhF at the cell poles that may otherwise lead to hyperflagellation, likely by antagonizing FleQ-dependent transcriptional activation. FimV is essential to FleN polar location. FimV and FleN temporally regulate the onset of flagellar assembly by preventing premature polar targeting of FlhF and the ensuing premature targeting of additional flagellar components. Our results shed new light on the mechanisms that ensure the timely assembly of the appropriate number of flagella at the correct polar location in polarly flagellated bacteria. HIGHLIGHTSO_LIFimV, FlhF and FleN determine the position, number and timing of flagellar assembly C_LIO_LIFimV is essential to the normal intracellular distribution of FlhF and FleN C_LIO_LIFlhF restricts flagellar location to the cell poles and promotes efficient assembly C_LIO_LISoluble, cytoplasmic FleN prevents polar FlhF accumulation and hyperflagellation C_LIO_LIPole-bound FimV and FleN prevent premature FlhF recruitment and flagellar assembly C_LI GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=121 SRC="FIGDIR/small/571843v5_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@f57b6dorg.highwire.dtl.DTLVardef@7a38c7org.highwire.dtl.DTLVardef@259c58org.highwire.dtl.DTLVardef@13b6f20_HPS_FORMAT_FIGEXP M_FIG C_FIG