The
            pilH
            gene encodes an ABC transporter homologue required for type IV pilus biogenesis and social gliding motility in
            Myxococcus xanthus

ABSTRACT In Myxococcus xanthus, two-component systems have crucial roles in regulating motility behavior and development. Here we describe an orphan response regulator, consisting of an N-terminal receiver domain and a C-terminal DNA binding domain, which is required for A and type IV pilus-dependent gliding motility. Genetic evidence suggests that phosphorylation of the conserved, phosphorylatable aspartate residue in the receiver domain is required for DigR activity. Consistent with the defect in type IV pilus-dependent motility, a digR mutant is slightly reduced in type IV pilus biosynthesis, and the composition of the extracellular matrix fibrils is abnormal, with an increased content of polysaccharides and decreased accumulation of the FibA metalloprotease. By using genome-wide transcriptional profiling, 118 genes were identified that are directly or indirectly regulated by DigR. These 118 genes include only 2, agmQ and cheY4, previously implicated in A and type IV pilus-dependent motility, respectively. In silico analyses showed that 36% of the differentially expressed genes are likely to encode exported proteins. Moreover, four genes encoding homologs of extracytoplasmic function (ECF) sigma factors, which typically control aspects of cell envelope homeostasis, are differentially expressed in a digR mutant. We suggest that the DigR response regulator has an important function in cell envelope homeostasis and that the motility defects in a digR mutant are instigated by the abnormal cell envelope and abnormal expression of agmQ and cheY4.

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Cyclic Di-GMP Regulates Type IV Pilus-Dependent Motility in Myxococcus xanthus

Journal of Bacteriology ◽

10.1128/jb.00281-15 ◽

2015 ◽

Vol 198 (1) ◽

pp. 77-90 ◽

Cited By ~ 26

Author(s):

Dorota Skotnicka ◽

Tobias Petters ◽

Jan Heering ◽

Michael Hoppert ◽

Volkhard Kaever ◽

...

Keyword(s):

Myxococcus Xanthus ◽

Lifestyle Changes ◽

Gliding Motility ◽

Cyclase Activity ◽

Diguanylate Cyclase ◽

Type Iv Pilus ◽

Cell Agglutination ◽

Content Type ◽

Type Iv ◽

Altered Cell

ABSTRACTThe nucleotide-based second messenger bis-(3′-5′)-cyclic dimeric GMP (c-di-GMP) is involved in regulating a plethora of processes in bacteria that are typically associated with lifestyle changes.Myxococcus xanthusundergoes major lifestyle changes in response to nutrient availability, with the formation of spreading colonies in the presence of nutrients and spore-filled fruiting bodies in the absence of nutrients. Here, we investigated the function of c-di-GMP inM. xanthusand show that this bacterium synthesizes c-di-GMP during growth. Manipulation of the c-di-GMP level by expression of either an active, heterologous diguanylate cyclase or an active, heterologous phosphodiesterase correlated with defects in type IV pilus (T4P)-dependent motility, whereas gliding motility was unaffected. An increased level of c-di-GMP correlated with reduced transcription of thepilAgene (which encodes the major pilin of T4P), reduced the assembly of T4P, and altered cell agglutination, whereas a decreased c-di-GMP level correlated with altered cell agglutination. The systematic inactivation of the 24 genes inM. xanthusencoding proteins containing GGDEF, EAL, or HD-GYP domains, which are associated with c-di-GMP synthesis, degradation, or binding, identified three genes encoding proteins important for T4P-dependent motility, whereas all mutants had normal gliding motility. Purified DmxA had diguanylate cyclase activity, whereas the hybrid histidine protein kinases TmoK and SgmT, each of which contains a GGDEF domain, did not have diguanylate cyclase activity. These results demonstrate that c-di-GMP is important for T4P-dependent motility inM. xanthus.IMPORTANCEWe provide the first direct evidence thatM. xanthussynthesizes c-di-GMP and demonstrate that c-di-GMP is important for T4P-dependent motility, whereas we did not obtain evidence that c-di-GMP regulates gliding motility. The data presented uncovered a novel mechanism for regulation of T4P-dependent motility, in which increased levels of c-di-GMP inhibit transcription of thepilAgene (which encodes the major pilin of T4P), ultimately resulting in the reduced assembly of T4P. Moreover, we identified an enzymatically active diguanylate cyclase that is important for T4P-dependent motility.

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The Myxococcus xanthus pilQ(sglA) Gene Encodes a Secretin Homolog Required for Type IV Pilus Biogenesis, Social Motility, and Development

Journal of Bacteriology ◽

10.1128/jb.181.1.24-33.1999 ◽

1999 ◽

Vol 181 (1) ◽

pp. 24-33 ◽

Cited By ~ 112

Author(s):

Daniel Wall ◽

Paul E. Kolenbrander ◽

Dale Kaiser

Keyword(s):

Outer Membrane ◽

Spontaneous Mutation ◽

Myxococcus Xanthus ◽

Type Iv Pili ◽

Gliding Motility ◽

Missense Mutations ◽

Fruiting Body Formation ◽

Subsequent Work ◽

Type Iv ◽

Pilus Biogenesis

ABSTRACT The Myxococcus xanthus sglA1 spontaneous mutation was originally isolated because it allowed dispersed cell growth in liquid yet retained the ability to form fruiting bodies. Consequently, most of today’s laboratory strains either contain the sglA1mutation or were derived from strains that carry it. Subsequent work showed that sglA was a gene for social gliding motility, a process which is mediated by type IV pili. Here sglA is shown to map to the major pil cluster and to encode a 901-amino-acid open reading frame (ORF) that is homologous to the secretin superfamily of proteins. Secretins form a channel in the outer membrane for the transport of macromolecules. The closest homologs found were PilQ proteins from Pseudomonas aeruginosa andNeisseria gonorrhoeae, which are required for type IV pili biogenesis and twitching motility. To signify these molecular and functional similarities, we have changed the name of sglAto pilQ. The hypomorphic pilQ1(sglA1) allele was sequenced and found to contain two missense mutations at residues 741 (G→S) and 762 (N→G). In addition, 19 independent social (S)-motility mutations are shown to map to the pilQ locus. In-frame deletions of pilQand its downstream gene, orfL, were constructed.pilQ is shown to be essential for pilus biogenesis, S-motility, rippling, and fruiting body formation, whileorfL is dispensable for these processes. ThepilQ1 allele, but not the ΔpilQ allele, was found to render cells hypersensitive to vancomycin, suggesting that PilQ1 alters the permeability properties of the outer membrane. Many differences between pilQ1 and pilQ +strains have been noted in the literature. We discuss some of these observations and how they may be rationalized in the context of our molecular and functional findings.

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Faculty Opinions recommendation of The Che4 pathway of Myxococcus xanthus regulates type IV pilus-mediated motility.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1019833.226437 ◽

2004 ◽

Author(s):

George Ordal

Keyword(s):

Myxococcus Xanthus ◽

Type Iv Pilus ◽

Type Iv

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Identification and initial characterization of a new pair of sibling sRNAs of Neisseria gonorrhoeae involved in type IV pilus biogenesis

Microbiology ◽

10.1099/mic.0.001080 ◽

2021 ◽

Vol 167 (9) ◽

Author(s):

Marie Zachary ◽

Susanne Bauer ◽

Maximilian Klepsch ◽

Katharina Wagler ◽

Bruno Hüttel ◽

...

Keyword(s):

Target Genes ◽

Target Prediction ◽

Type Iv Pilus ◽

Content Type ◽

Type Iv ◽

Gene Expression Control ◽

Initial Characterization ◽

Pilus Biogenesis ◽

Regulatory Rnas

Non-coding regulatory RNAs mediate post-transcriptional gene expression control by a variety of mechanisms relying mostly on base-pairing interactions with a target mRNA. Though a plethora of putative non-coding regulatory RNAs have been identified by global transcriptome analysis, knowledge about riboregulation in the pathogenic Neisseriae is still limited. Here we report the initial characterization of a pair of sRNAs of N. gonorrhoeae , TfpR1 and TfpR2, which exhibit a similar secondary structure and identical single-stranded seed regions, and therefore might be considered as sibling sRNAs. By combination of in silico target prediction and sRNA pulse expression followed by differential RNA sequencing we identified target genes of TfpR1 which are involved in type IV pilus biogenesis and DNA damage repair. We provide evidence that members of the TfpR1 regulon can also be targeted by the sibling TfpR2.

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Purification and Three-Dimensional Electron Microscopy Structure of the Neisseria meningitidis Type IV Pilus Biogenesis Protein PilG

Journal of Bacteriology ◽

10.1128/jb.00648-07 ◽

2007 ◽

Vol 189 (17) ◽

pp. 6389-6396 ◽

Cited By ~ 25

Author(s):

Richard F. Collins ◽

Muhammad Saleem ◽

Jeremy P. Derrick

Keyword(s):

Electron Microscopy ◽

Neisseria Meningitidis ◽

Metal Ion ◽

Polyacrylamide Gel Electrophoresis ◽

Three Dimensional ◽

Type Ii Secretion ◽

Type Ii ◽

Type Iv Pilus ◽

Type Iv ◽

Pilus Biogenesis

ABSTRACT Type IV pili are surface-exposed retractable fibers which play a key role in the pathogenesis of Neisseria meningitidis and other gram-negative pathogens. PilG is an integral inner membrane protein and a component of the type IV pilus biogenesis system. It is related by sequence to the extensive GspF family of secretory proteins, which are involved in type II secretion processes. PilG was overexpressed and purified from Escherichia coli membranes by detergent extraction and metal ion affinity chromatography. Analysis of the purified protein by perfluoro-octanoic acid polyacrylamide gel electrophoresis showed that PilG formed dimers and tetramers. A three-dimensional (3-D) electron microscopy structure of the PilG multimer was determined using single-particle averaging applied to samples visualized by negative staining. Symmetry analysis of the unsymmetrized 3-D volume provided further evidence that the PilG multimer is a tetramer. The reconstruction also revealed an asymmetric bilobed structure approximately 125 Å in length and 80 Å in width. The larger lobe within the structure was identified as the N terminus by location of Ni-nitrilotriacetic acid nanogold particles to the N-terminal polyhistidine tag. We propose that the smaller lobe corresponds to the periplasmic domain of the protein, with the narrower “waist” region being the transmembrane section. This constitutes the first report of a 3-D structure of a member of the GspF family and suggests a physical basis for the role of the protein in linking cytoplasmic and periplasmic protein components of the type II secretion and type IV pilus biogenesis systems.

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Three-dimensional observations of an aperiodic oscillatory gliding motility behaviour in Myxococcus xanthus using confocal interference reflection microscopy

10.1101/722231 ◽

2019 ◽

Author(s):

Liam M. Rooney ◽

Lisa S. Kölln ◽

Ross Scrimgeour ◽

William B. Amos ◽

Paul A. Hoskisson ◽

...

Keyword(s):

Myxococcus Xanthus ◽

Three Dimensional ◽

Basal Membrane ◽

Gliding Motility ◽

The Novel ◽

Force Transmission ◽

Topological Information ◽

Type Iv ◽

Microscopy Techniques

The Delta-proteobacterium, Myxococcus xanthus, has been used as a model for bacterial motility and to provide insights of bacterial swarming behaviours. Fluorescence microscopy techniques have shown that various mechanisms are involved in gliding motility, but these have almost entirely been limited to 2D studies and there is currently no understanding of gliding motility in a 3D context. We present here the first use of confocal interference reflection microscopy (IRM) to study gliding bacteria, and we reveal aperiodic oscillatory behaviour with changes in the position of the basal membrane relative to the coverglass on the order of 90 nm in vitro. Firstly, we use a model plano-convex lens specimen to show how topological information can be obtained from the wavelength-dependent interference pattern in IRM. We then use IRM to observe gliding M. xanthus and show that cells undergo previously unobserved changes in their height as they glide. We compare the wild-type with mutants of reduced motility, which also exhibit the same changes in adhesion profile during gliding. We find that the general gliding behaviour is independent of the proton motive force-generating complex, AglRQS, and suggest that the novel behaviour we present here may be a result of recoil and force transmission along the length of the cell body following firing of the Type IV pili.

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Analysis of type IV pilus and its associated motility in Myxococcus xanthus using an antibody reactive with native pilin and pili

Microbiology ◽

10.1099/mic.0.27614-0 ◽

2005 ◽

Vol 151 (2) ◽

pp. 353-360 ◽

Cited By ~ 23

Author(s):

Yinuo Li ◽

Renate Lux ◽

Andrew E. Pelling ◽

James K. Gimzewski ◽

Wenyuan Shi

Keyword(s):

Amino Acids ◽

Current Model ◽

Bacterial Species ◽

Myxococcus Xanthus ◽

Type Iv Pili ◽

Gliding Motility ◽

Soluble Form ◽

Truncated Form ◽

Type Iv ◽

Oligomerization Domain

Myxococcus xanthus possesses a social gliding motility that requires type IV pili (TFP). According to the current model, M. xanthus pili attach to an external substrate and retract, pulling the cell body forward along their long axis. By analogy with the situation in other bacteria employing TFP-dependent motility, M. xanthus pili have been assumed to be composed of pilin (PilA) subunits, but this has not previously been confirmed. The first 28 amino acids of the M. xanthus PilA protein share extensive homology with the N-terminal oligomerization domain of pilins in other bacterial species. To facilitate purification, the authors engineered a truncated form of M. xanthus PilA lacking the first 28 amino acids and purified this protein in soluble form. Polyclonal antibody generated against this protein was reactive with native pilin and pili. Using this antibody, it was confirmed that TFP of M. xanthus are indeed composed of PilA, and that TFP are located unipolarly and required for social gliding motility via retraction. Using tethering as well as motility assays, details of pili function in M. xanthus social motility were further examined.

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