Biology subjects

Sogaard-Andersen, L.

Publications and source records attributed to Sogaard-Andersen, L..

5 recordsLinked to original sources

bioRxiv2021

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

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. Importancec-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 upstream of 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 downstream of the Frz system to regulate motility. Together with previous observations, we conclude that PilZ-domain proteins and c-di-GMP act in multiple parallel pathways to regulate motility and development in M. xanthus.

microbiology↗

bioRxiv2020

PomX, a ParA/MinD ATPase activating protein, is a triple regulator of cell division in Myxococcus xanthus

Cell division is precisely regulated to generate daughter cells of correct size and shape. In the social bacterium Myxococcus xanthus, the tripartite PomX/Y/Z complex directly stimulates positioning of the cytokinetic FtsZ-ring at midcell to mark the division site. The [~]15 MDa PomX/Y/Z complex associates with the nucleoid in a PomZ-dependent manner, translocates to midcell to stimulate FtsZ-ring formation, and undergoes fission during division. We demonstrate that PomX consists of two functionally distinct domains and has three functions. The N-terminal domain interacts with the ParA/MinD ATPase PomZ and stimulates PomZ ATPase activity. The C-terminal domain mediates PomX self-interaction, interaction to PomY, and serves as a scaffold for PomX/Y/Z complex formation. Moreover, the PomX/PomZ interaction is important for fission. These observations together with previous work support that the architecturally diverse ATPase activating proteins of ParA/MinD ATPases are highly modular and use the same mechanism to activate their cognate ATPase via a short positively charged N-terminal extension.

microbiology↗

bioRxiv2020

SMC and the bactofilin/PadC scaffold have distinct yet redundant functions in chromosome segregation and organization in Myxococcus xanthus

In bacteria, ParABS systems and structural maintenance of chromosome (SMC) condensin-like complexes are important for chromosome segregation and organization. The rod-shaped Myxococcus xanthus cells have a unique chromosome arrangement in which a scaffold composed of three bactofilins (BacNOP) and PadC positions the essential ParB{middle dot}parS segregation complexes and the DNA segregation ATPase ParA in the subpolar regions. Here, we identify the Smc and ScpAB subunits of the SMC complex in M. xanthus and demonstrate that SMC is conditionally essential with mutants containing smc or scpAB deletions being temperature sensitive. Lack of SMC caused defects in chromosome segregation and organization. Lack of the BacNOP/PadC scaffold caused chromosome segregation defects but was not essential. Inactivation of SMC was synthetic lethal with lack of the BacNOP/PadC scaffold. Lack of SMC interfered with formation of the BacNOP/PadC scaffold while lack of this scaffold did not interfere with chromosome association by SMC. Altogether, our data support that three systems cooperate to enable chromosome segregation in M. xanthus, whereby ParABS constitutes the basic machinery and SMC and the BacNOP/PadC scaffold have distinct yet redundant roles in this process with SMC supporting individualization of daughter chromosomes and BacNOP/PadC making the ParABS system operate more robustly

microbiology↗

bioRxiv2020

Four different mechanisms for switching cell polarity

The mechanisms and design principles of regulatory systems establishing stable polarized protein patterns within cells are well studied. However, cells can also dynamically control their cell polarity. Here, we ask how an upstream signaling system can switch the orientation of a polarized pattern. We use a mathematical model of a core polarity system based on three proteins as the basis to study different mechanisms of signal-induced polarity switching. The analysis of this model reveals four general classes of switching mechanisms with qualitatively distinct behaviors: the transient oscillator switch, the reset switch, the prime-release switch, and the push switch. Each of these regulatory mechanisms effectively implements the function of a spatial toggle switch, however with different characteristics in their nonlinear and stochastic dynamics. We identify these characteristics and also discuss experimental signatures of each type of switching mechanism.

biophysics↗

bioRxiv2020

Characterization of the exopolysaccharide biosynthesis pathway in Myxococcus xanthus

Myxococcus 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.

microbiology↗