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

1999 ◽  
Vol 181 (1) ◽  
pp. 24-33 ◽  
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.

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

Microbiology ◽  
2005 ◽  
Vol 151 (2) ◽  
pp. 353-360 ◽  
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.

scholarly journals Phosphorylation and Dephosphorylation among Dif Chemosensory Proteins Essential for Exopolysaccharide Regulation in Myxococcus xanthus

2010 ◽  
Vol 192 (17) ◽  
pp. 4267-4274 ◽  
Author(s):  
Wesley P. Black ◽  
Florian D. Schubot ◽  
Zhuo Li ◽  
Zhaomin Yang
Keyword(s):  
Double Mutant ◽  
Kinase Activity ◽  
Myxococcus Xanthus ◽  
Type Iv Pili ◽  
Gliding Motility ◽  
Chemosensory Proteins ◽  
Downstream Target ◽  
Signaling Complex ◽  
Type Iv

ABSTRACT Myxococcus xanthus social gliding motility, which is powered by type IV pili, requires the presence of exopolysaccharides (EPS) on the cell surface. The Dif chemosensory system is essential for the regulation of EPS production. It was demonstrated previously that DifA (methyl-accepting chemotaxis protein [MCP]-like), DifC (CheW-like), and DifE (CheA-like) stimulate whereas DifD (CheY-like) and DifG (CheC-like) inhibit EPS production. DifD was found not to function downstream of DifE in EPS regulation, as a difD difE double mutant phenocopied the difE single mutant. It has been proposed that DifA, DifC, and DifE form a ternary signaling complex that positively regulates EPS production through the kinase activity of DifE. DifD was proposed as a phosphate sink of phosphorylated DifE (DifE∼P), while DifG would augment the function of DifD as a phosphatase of phosphorylated DifD (DifD∼P). Here we report in vitro phosphorylation studies with all the Dif chemosensory proteins that were expressed and purified from Escherichia coli. DifE was demonstrated to be an autokinase. Consistent with the formation of a DifA-DifC-DifE complex, DifA and DifC together, but not individually, were found to influence DifE autophosphorylation. DifD, which did not inhibit DifE autophosphorylation directly, was found to accept phosphate from autophosphorylated DifE. While DifD∼P has an unusually long half-life for dephosphorylation in vitro, DifG efficiently dephosphorylated DifD∼P as a phosphatase. These results support a model where DifE complexes with DifA and DifC to regulate EPS production through phosphorylation of a downstream target, while DifD and DifG function synergistically to divert phosphates away from DifE∼P.

scholarly journals Myxococcus xanthus dif Genes Are Required for Biogenesis of Cell Surface Fibrils Essential for Social Gliding Motility

2000 ◽  
Vol 182 (20) ◽  
pp. 5793-5798 ◽  
Author(s):  
Zhaomin Yang ◽  
Xiaoyuan Ma ◽  
Leming Tong ◽  
Heidi B. Kaplan ◽  
Lawrence J. Shimkets ◽  
...  
Keyword(s):  
Cell Surface ◽  
Myxococcus Xanthus ◽  
Bacterial Chemotaxis ◽  
Type Iv Pili ◽  
Gliding Motility ◽  
Genetic Studies ◽  
Developmental Defects ◽  
Bacterial Surface ◽  
Type Iv ◽  
Cell Surface Structures

ABSTRACT Myxococcus xanthus social (S) gliding motility has been previously reported by us to require the chemotaxis homologues encoded by the dif genes. In addition, two cell surface structures, type IV pili and extracellular matrix fibrils, are also critical to M. xanthus S motility. We have demonstrated here that M. xanthus dif genes are required for the biogenesis of fibrils but not for that of type IV pili. Furthermore, the developmental defects of dif mutants can be partially rescued by the addition of isolated fibril materials. Along with the chemotaxis genes of various swarming bacteria and the pilGHIJ genes of the twitching bacteriumPseudomonas aeruginosa, the M. xanthus dif genes belong to a unique class of bacterial chemotaxis genes or homologues implicated in the biogenesis of structures required for bacterial surface locomotion. Genetic studies indicate that the dif genes are linked to theM. xanthus dsp region, a locus known to be crucial forM. xanthus fibril biogenesis and S gliding.

scholarly journals Characterization of the exopolysaccharide biosynthesis pathway in Myxococcus xanthus

2020 ◽  
Author(s):  
María Pérez-Burgos ◽  
Inmaculada García-Romero ◽  
Jana Jung ◽  
Eugenia Schander ◽  
Miguel A. Valvano ◽  
...  
Keyword(s):  
Fruiting Body ◽  
Repeat Unit ◽  
Myxococcus Xanthus ◽  
Type Iv Pili ◽  
Fruiting Body Formation ◽  
Body Formation ◽  
Polysaccharide Biosynthesis ◽  
Type Iv ◽  
Biosynthetic Machinery ◽  
Dependent Pathway

AbstractMyxococcus xanthus arranges into two morphologically distinct biofilms depending on its nutritional status, i.e. coordinately spreading colonies in the presence of nutrients and spore-filled fruiting bodies in the absence of nutrients. A secreted polysaccharide referred to as exopolysaccharide (EPS) is a structural component of both biofilms and is also important for type IV pili-dependent motility and fruiting body formation. Here, we characterize the biosynthetic machinery responsible for EPS biosynthesis using bioinformatics, genetics, heterologous expression, and biochemical experiments. We show that this machinery constitutes a Wzx/Wzy-dependent pathway dedicated to EPS biosynthesis. Our data support that EpsZ (MXAN_7415) is the polyisoprenyl-phosphate hexose-1-phosphate transferase responsible for initiation of the repeat unit synthesis. Heterologous expression experiments support that EpsZ has galactose-1-P transferase activity. Moreover, MXAN_7416, renamed WzxEPS, and MXAN_7442, renamed WzyEPS, are the Wzx flippase and Wzy polymerase responsible for translocation and polymerization of the EPS repeat unit, respectively. Also, in this pathway, EpsV (MXAN_7421) is the polysaccharide co-polymerase and EpsY (MXAN_7417) the outer membrane polysaccharide export (OPX) protein. Mutants with single in-frame deletions in the five corresponding genes had defects in type IV pili-dependent motility and a conditional defect in fruiting body formation. Furthermore, all five mutants were deficient in type IV pili formation and genetic analyses suggest that EPS and/or the EPS biosynthetic machinery stimulates type IV pili extension. Additionally, we identify a polysaccharide biosynthesis gene cluster, which together with an orphan gene encoding an OPX protein make up a complete Wzx/Wzy-dependent pathway for synthesis of an unknown polysaccharide.ImportanceThe secreted polysaccharide referred to as exopolysaccharide (EPS) has important functions in the social life cycle of M. xanthus; however, little is known about how EPS is synthesized. Here, we characterized the EPS biosynthetic machinery and show that it makes up a Wzx/Wzy-dependent pathway for polysaccharide biosynthesis. Mutants lacking a component of this pathway had reduced type IV pili-dependent motility and a conditional defect in development. Also, these analysis suggest that EPS and/or the EPS biosynthetic machinery is important for type IV pili formation.

scholarly journals Three PilZ domain proteins, PlpA, PixA and PixB, have distinct functions in regulation of motility and development in Myxococcus xanthus

2021 ◽  
Author(s):  
Sofya Kuzmich ◽  
Dorota Skotnicka ◽  
Dobromir Szadkowski ◽  
Philipp Klos ◽  
María Pérez‐Burgos ◽  
...  
Keyword(s):  
Bacterial Species ◽  
Myxococcus Xanthus ◽  
Type Iv Pili ◽  
Gliding Motility ◽  
Intact Protein ◽  
Dependent Manner ◽  
Chemosensory System ◽  
Type Iv ◽  
Acetyltransferase Domain

In bacteria, the nucleotide-based second messenger bis-(3’-5’)-cyclic dimeric GMP (c-di-GMP) binds to effectors to generate outputs in response to changes in the environment. In Myxococcus xanthus, c-di-GMP regulates type IV pili-dependent motility and the starvation-induced developmental program that results in formation of spore-filled fruiting bodies; however, little is known about the effectors that bind c-di-GMP. Here, we systematically inactivated all 24 genes encoding PilZ domain-containing proteins, which are among the most common c-di-GMP effectors. We confirm that the stand-alone PilZ-domain protein PlpA is important for regulation of motility independently of the Frz chemosensory system, and that Pkn1, which is composed of a Ser/Thr kinase domain and a PilZ domain, is specifically important for development. Moreover, we identify two PilZ-domain proteins that have distinct functions in regulating motility and development. PixB, which is composed of two PilZ domains and an acetyltransferase domain, binds c-di-GMP in vitro and regulates type IV pili-dependent and gliding motility in a Frz-dependent manner as well as development. The acetyltransferase domain is required and sufficient for function during growth while all three domains and c-di-GMP binding are essential for PixB function during development. PixA is a response regulator composed of a PilZ domain and a receiver domain, binds c-di-GMP in vitro, and regulates motility independently of the Frz system likely by setting up the polarity of the two motility systems. Our results support a model whereby PlpA, PixA and PixB act in independent pathways and have distinct functions in regulation of motility. Importance c-di-GMP signaling controls bacterial motility in many bacterial species by binding to downstream effector proteins. Here, we identify two PilZ domain-containing proteins in Myxococcus xanthus that bind c-di-GMP. We show that PixB, which contains two PilZ domains and an acetyltransferase domain, acts in a manner that depends on the Frz chemosensory system to regulate motility via the acetyltransferase domain while the intact protein and c-di-GMP binding are essential for PixB to support development. By contrast, PixA acts acts in Frz-independent mannerto regulate motility. Together with previous observations, we conclude that PilZ-domain proteins and c-di-GMP act in multiple independent pathways to regulate motility and development in M. xanthus.

scholarly journals Three PilZ domain proteins, PlpA, PixA and PixB, have distinct functions in regulation of motility and development in Myxococcus xanthus

2021 ◽  
Author(s):  
Sofya Kuzmich ◽  
Dorota Skotnicka ◽  
Dobromir Szadkowski ◽  
Philipp Klos ◽  
Maria Perez-Burgos ◽  
...  
Keyword(s):  
Response Regulator ◽  
Myxococcus Xanthus ◽  
Type Iv Pili ◽  
Gliding Motility ◽  
Chemosensory System ◽  
Developmental Program ◽  
Type Iv ◽  
Genes Encoding ◽  
Acetyltransferase Domain

In bacteria, the nucleotide-based second messenger bis-(3'-5')-cyclic dimeric GMP (c-di-GMP) binds to effectors to generate outputs in response to changes in the environment. In Myxococcus xanthus, c-di-GMP regulates type IV pili-dependent motility and the starvation-induced developmental program that results in the formation of spore-filled fruiting bodies; however, little is known about the effectors that bind c-di-GMP. Here, we systematically inactivated all 24 genes encoding PilZ domain-containing proteins, which are among the most common c-di-GMP receptors. We confirm that PlpA, a stand-alone PilZ-domain protein, is specifically important for motility and that Pkn1, which is composed of a Ser/Thr domain and a PilZ domain, is specifically important for development. Moreover, we identify two PilZ-domain proteins that have distinct functions in regulating motility and development. PixB, which is composed of two PilZ domains and an acetyltransferase domain, binds c-di-GMP in vitro and regulates type IV pili-dependent and gliding motility upstream of the Frz chemosensory system as well as development. The acetyltransferase domain is required and sufficient for function during growth while all three domains and c-di-GMP binding are essential for PixB function during development. PixA is a response regulator composed of a PilZ domain and a receiver domain, binds c-di-GMP in vitro, and regulates motility downstream of the Frz chemosensory system by setting up the polarity of the two motility systems. Our results support a model whereby the three proteins PlpA, PixA and PixB act in parallel pathways and have distinct functions to regulation of motility.

Type IV pili‐dependent gliding motility in the Gram‐positive pathogenClostridium perfringensand other Clostridia

2006 ◽  
Vol 62 (3) ◽  
pp. 680-694 ◽  
Author(s):  
John J. Varga ◽  
Van Nguyen ◽  
David K. O'Brien ◽  
Katherine Rodgers ◽  
Richard A. Walker ◽  
...  
Keyword(s):  
Type Iv Pili ◽  
Gliding Motility ◽  
Gram Positive ◽  
Type Iv

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

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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