Dynamic localization of HmpF regulates type IV pilus activity and directional motility in the filamentous cyanobacterium Nostoc punctiforme

Abstract

Many cyanobacteria exhibit surface motility powered by type 4 pili (T4P). In the model filamentous cyanobacterium Nostoc punctiforme, the T4P systems are arrayed in static, bipolar rings in each cell. The chemotaxis-like Hmp system is essential for motility and the coordinated polar accumulation of PilA on cells in motile filaments, while the Ptx system controls positive phototaxis. Using transposon mutagenesis, a gene, designated hmpF, was identified as involved in motility. Synteny among filamentous cyanobacteria and the similar expression patterns for hmpF and hmpD imply that HmpF is part of the Hmp system. Deletion of hmpF produced a phenotype distinct from other hmp genes, but indistinguishable from pilB or pilQ. Both an HmpF-GFPuv fusion protein, and PilA, as assessed by in situ immunofluorescence, displayed coordinated, unipolar localization at the leading pole of each cell. Reversals were modulated by changes in light intensity and preceded by the migration of HmpF-GFPuv to the lagging cell poles. These results are consistent with a model where direct interaction between HmpF and the T4P system activates pilus extension, the Hmp system facilitates coordinated polarity of HmpF to establish motility, and the Ptx system modulates HmpF localization to initiate reversals in response to changes in light intensity.

A partner‐switching regulatory system controls hormogonium development in the filamentous cyanobacterium Nostoc punctiforme

“…To provide additional evidence supporting the hypothesis that these three proteins comprise the cognate components of a PSRS, a comparative genomics approach was taken, using a previously generated data set (Cho et al, ) to determine the co‐occurrence of orthologs for each protein in the predicted proteome of 125 cyanobacteria with sequenced genomes (Fig. B).…”

“…Preparation of cell material, protein extraction and detection of PilA, RbcL, HmpD and GFPuv by immunoblot analysis were performed as previously described (Cho et al, ). Preparation, detection and quantification of soluble HPS in the culture medium by lectin blotting with biotinylated UEA (Vector Laboratories) were performed as previously described (Khayatan et al, ).…”

“…Simultaneous detection of PilA and HPS by immunofluorescence and fluorescent lectin staining was performed as previously described (Cho et al, ). Quantification of PilA‐derived immunofluorescence was performed in the same manner as that described for UEA‐fluorescein staining (Cho et al, ).…”

“…To test this hypothesis, RT-qPCR was employed to quantify the expression of several genes whose products are required for normal PilA and HPS secretion (Fig. 5), including pilB, pilQ, hpsE and the recently characterized hmpF (Risser and Meeks, 2013;Khayatan et al, 2015;Cho et al, 2017). Prior to hormogonium induction, expression levels for each gene were similar in the wildtype and ΔhmpV strain, with a slight reduction in expression for several genes in the ΔhmpV strain that was not GFPuv-derived fluorescence (cyan) for the wild type, and strains harboring gfpuv-tagged alleles of Hmp PSRS genes (as indicated), 0 and 24 h after hormogonium induction (as indicated).…”

“…Together, pilA and a collection of other genes from the pil and hps loci are thought to produce components of the T4P system that drives gliding motility and the secretion of HPS in hormogonia (Khayatan et al, ). N. punctiforme also displays positive phototaxis, which is regulated by genes encoded at the ptx locus (Campbell et al, ), and this directional motility is thought to be controlled by the recently characterized protein HmpF, which dynamically localizes to the leading poles in motile hormogonia and is essential for activation of T4P motors (Cho et al, ).…”