Exopolysaccharide biosynthesis genes required for social motility in Myxococcus xanthus

Abstract

Summary Social (S)-motility inMyxococcus xanthus is a flagellum-independent gliding motility system that allows bacteria to move in groups on solid surfaces. S-motility has been shown to require type IV pili (TFP), exopolysaccharide (EPS; a component of fibrils) and lipopolysaccharide (LPS). Previously, information concerning EPS biogenesis in M. xanthus was lacking. In this study, we screened 5000 randomly mutagenized colonies for defects in S-motility and EPS and identified two genetic regions essential for EPS biogenesis: the EPS synthesis ( eps ) region and the EPS -associated ( eas ) region. Mutants with insertions in the eps and eas regions were defective in Smotility and fruiting body formation. These mutants failed to bind the dye calcofluor white, indicating that they lacked EPS; however, they retained normal TFP and LPS. Analysis of the eps locus showed several open reading frames (ORFs) that encode homologues to glycosyltransferases, glucanases and EPS transporters as well as regulatory proteins; the eas locus contains two ORFs: one exhibits homology to hypothetical proteins with a conserved domain of unknown function and the other displays no apparent homology to other proteins in the database. Further genetic mutagenesis analysis indicates that the whole eps region is involved in the biosynthesis of fibrils and fibril EPS. The operon at the proximal end of the eps region was analysed by generating in-frame deletion mutations. These mutants showed varying degrees of defects in the bacterium's ability to produce EPS or perform EPS-related functions, confirming the involvement of these genes in M. xanthus EPS biogenesis.

Synthesis and Assembly of a Novel Glycan Layer in Myxococcus xanthus Spores

“…6, step 2) (35). Two flippase homologs can be identified in the M. xanthus genome, MXAN_1035 and MXAN_7416, which are located in the vicinity of polysaccharide synthesis protein homologs previously implicated in EPS production (32,47). Based on the observation that deletion of MXAN_1035 had only a minor effect on sporulation, the most likely candidate is MXAN_7416.…”

“…Previous studies have demonstrated that almost all of the carbohydrate subunits identified in our analyses have been shown to be part of at least one of the two other cell surface polysaccharides of M. xanthus as follows: the EPS and the O-antigen (O-Ag) component of the lipopolysaccharide (LPS) (29,30). To determine whether O-Ag or EPS plays an essential role as a source of the spore coat components, or whether these components arise as a contamination due to co-purification, we generated a double mutant bearing disruptions in genes that have previously been shown to be necessary for O-antigen (wzm) (31) and EPS (epsV) synthesis (32). The wzm epsV double mutant formed as many heat-and sonication-resistant spores as the wild type (163 Ϯ 37% versus 100 Ϯ 20%, respectively) suggesting that neither of these products are essential for formation of resistant spores.…”

Inorganic polyphosphate in the social life of Myxococcus xanthus : Motility, development, and predation

“…The PPK1 mutant, which is strikingly defective in S-motility, accumulates an excessive amount of the cell-surface pilin; this hyperpiliation phenotype is likely due to an insufficient amount of fibril EPS (27) and perhaps other fibril components. Biogenesis of EPS involves genes eps and eas (35), a DnaK homolog (encoded by sglK) (36), chemotaxis homologues (dif ) genes (37), and some uncharacterized dsp genes. EPS-deficient mutants fail to bind calcofluor white dye (38) and are defective in cellular agglutination, S-motility, and fruiting body formation (25,26), phenotypes similar to those in the PPK1 mutant.…”

Transposon Insertions ofmagellan-4That Impair Social Gliding Motility inMyxococcus xanthus

“…General searches for mutants of M. xanthus with motility defects have shown that single-cell (adventurous, A) motility and group-dependent (social, S) motility are genetically separable and that each type of motility requires the functions of large, nonoverlapping sets of genes (Hodgkin and Kaiser 1979a,b;Macneil et al 1994a,b;Youderian et al 2003). Mutations in only three genes, mglA, which encodes a small GTPase (Stephens and Kaiser 1987;Stephens et al 1989;Hartzell and Kaiser 1991a,b;Hartzell 1997), agmA, which is predicted to encode a critical amidase involved in cell wall biogenesis (Youderian et al 2003), and epsI/nla24, predicted to encode an activator of transcription (Caberoy et al 2003;Lancero et al 2004;Lu et al 2005), have been shown to abolish both mechanisms of motility simultaneously.…”

“…1 The EPS component, but not the protein component, of fibrils appears to be essential for social motility and cell-cell adhesion. Mutants defective in EPS production are defective in S motility (Lu et al 2005), and EPS stimulates pilus retraction (Li et al 2003).…”