Dynamic proton-dependent motors power Type IX secretion and gliding adhesin movement inFlavobacterium
Maxence S. VincentCaterina Comas HervadaCorinne Sebban‐KreuzerHugo Le GuennoMaïalène ChabalierArtémis KostaFrançoise GuerlesquinTâm MignotMark J. McBrideEric CascalèsThierry Doan
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Abstract:
Abstract Motile bacteria usually rely on external apparatus like flagella for swimming or pili for twitching. By contrast, gliding bacteria do not rely on obvious surface appendages to move on solid surfaces. Flavobacterium johnsoniae and other bacteria in the Bacteroidetes phylum use adhesins whose movement on the cell surface supports motility. In F. johnsoniae , secretion and helicoidal motion of the main adhesin SprB are intimately linked and depend on the type IX secretion system (T9SS). Both processes necessitate the proton motive force (PMF), which is thought to fuel a molecular motor that comprises the GldL and GldM cytoplasmic membrane proteins. Here we show that F. johnsoniae gliding motility is powered by the pH gradient component of the PMF. We further delineate the interaction network between the GldLM transmembrane helices (TMH) and show that conserved glutamate residues in GldL TMH are essential for gliding motility, although having distinct roles in SprB secretion and motion. We then demonstrate that the PMF and GldL trigger conformational changes in the GldM periplasmic domain. We finally show that multiple GldLM complexes are distributed in the membrane suggesting that a network of motors may be present to move SprB along a helical path on the cell surface. Altogether, our results provide evidence that GldL and GldM assemble dynamic membrane channels that use the proton gradient to power both T9SS-dependent secretion of SprB and its motion at the cell surface.Keywords:
Gliding motility
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Periplasmic flagella are complex nanomachines responsible for distinctive morphology and motility of spirochetes. Although bacterial flagella have been extensively studied for several decades in the model systems Escherichia coli and Salmonella enterica, our understanding of periplasmic flagella in many disease-causing spirochetes remains incomplete. Recent advances, including molecular genetics, biochemistry, structural biology, and cryo-electron tomography, have greatly increased our understanding of structure and function of periplasmic flagella. In this chapter, we summarize some of the recent findings that provide new insights into the structure, assembly, and function of periplasmic flagella.
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Uropathogenic Escherichia coli are major causative agents of cystitis and pyelonephritis. Most E. coli pyelonephritis isolates express pili encoded by the pyelonephritis-associated pili (pap) gene cluster. The pap DNA sequence encodes pilin monomers that are assembled into pili fibers; pap also encodes adhesins that recent results suggest might be located at the tips of the pilus fibers. The study presented here is a status report of work that has two major goals: to determine (1) if any Pap proteins associate with pili and (2) if Pap proteins not required for pili assembly affect levels of pili-cell surface expression. To address the first aim, antisera to pili were used to precipitate pili from detergent extracts containing 35S-labeled Pap proteins. The results suggested that a protein of 16-kilodaltons apparent molecular mass associated with pili. Other interpretations of the data are discussed. The second aim was addressed by constructing E. coli strains that contained different pap regions. With the use of electron microscopy and a pili ELISA, it was found that E. coli containing a 6.5-kilobase-pair region of pap expressed low levels of pili, but no P-adhesin was detected. Transformation of this E. coli strain with a plasmid containing an additional 3.5-kilobase-pair pap DNA sequence resulted in an eightfold increase in pili expression as well as expression of P adhesin. These results indicated that pili expression was affected by Pap proteins not required for pili assembly.
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The interaction of differentially piliated Aeromonas strains expressing pili of two broadly different morphologic types (short, rigid (S/R) and/or long, wavy (L/W)) with human peripheral blood mononuclear leukocytes (PMN) was investigated to determine whether host defense cells might exert a selective pressure on pili expression in vivo accounting for the different pili phenotypes of clinical and environmental strains. A majority of Aeromonas veronii biotype sobria strains from water (6/6) and faeces (8/11) readily associated with PMN (> 60% PMN with adherent and/or internalised bacteria), irrespective of their degree, or predominant type, of piliation. Rigid pili of Aeromonas species did not promote interaction with PMN. However, the majority (55%) of strains which interacted well with PMN were adherent to HEp-2 cells. Interaction with PMN is unlikely to be the reason few S/R pili are seen on faecal strains, but it may be a selective pressure on L/W adhesive pili, or other OMP adhesins, resulting in the shedding of strains which have lost critical adhesins.
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Surface attachment of bacteria is the first step of biofilm formation and biofilms are associated with infections and bacterial resistance. Surface attachment of bacteria is often mediated by extracellular appendages, for example flagellum and pili. The flagellum is a cork-screw like structure used for swimming and surface sensing. Pili are filamentous structures and have a wide variety of functions, among them attachment on surfaces. Because of the small diameter of flagellum and pili, direct observations of flagellum and pili are challenging under physiological conditions.
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Although spirochete periplasmic flagella have many features similar to typical bacterial flagella, they are unique in their structure and internal periplasmic location. This location provides advantages for pathogenic spirochetes to enter and to adapt in the appropriate host, and to penetrate through matrices that inhibit the motility of most other bacteria. These flagella are complex, and they dynamically interact with the spirochete cell cylinder in novel ways. Electron microscopy, tomography and three-dimensional reconstructions have provided new insights into flagellar structure and its relationship to the spirochetal cell cylinder. Recent advances in genetic methods have begun to shed light on the composition of the spirochete flagellum, and on the regulation of its synthesis. Because spirochetes have a high length to width ratio, their cells provide an opportunity to study two important features. These include the polarity or distribution of flagellar synthesis as well as the mechanisms required for coordination of the movement of the cell ends, to enable it to move in the forward or reverse direction.
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