How myxobacteria regulate social behaviors by outer membrane exchange.

摘要

Myxococcus xanthus is an ideal organism to study social behavior. Development, predation and rippling are all hallmarks of social behaviors in myxobacteria. Our lab has uncovered outer membrane exchange (OME), which is one mechanism used by myxobacteria to cooperate and communicate in social environments. Two key players, TraA and TraB, were identified to be required for OME. TraA is a homophilic cell surface receptor that recognizes similar or identical TraA receptors on other cells leading to fusion of membranes and ultimately cell content exchange. The PA14-like domain in TraA is highly polymorphic and was found to be the molecular determinant of specificity, as similar TraA proteins will lead to exchange and dissimilar TraA proteins will not. One function of OME is to modulate development and motility behaviors. In this regard OME can cause 'swarm inhibition' where the motility of motile cells can be blocked when mixed with nonmotile cells. 'Relief of swarm inhibition' is observed when either the motile or nonmotile cells have a traA mutation.;Using swarm inhibition as a phenotype, a forward genetic screen was devised to isolate mutants involved in OME and in the downstream response that leads to swarm inhibition. Over 50 mutants defective in outer membrane exchange were identified indicating that the screen was effective. Mapping all these mutations revealed that the insertions were in TraA and TraB suggesting that there are few, if any, proteins besides TraA and TraB involved in the process of lipoprotein exchange. However, a new mutant class which is not defective in outer membrane exchange, but instead defective in the downstream process leading to swarm inhibition was identified. The mutation was mapped to MXAN_4426 and named outer membrane response protein A (OmrA). Through bioinformatic analysis a second gene was identified that when mutated resulted in a partial relief of swarm inhibition. The mutant was named OmrB. Interestingly, the two domains of these proteins are homologous to the MprF protein that functions in conferring resistance to the cationic antibiotic Daptomycin in Staphylococcus aureus. The mechanism of resistance involves changing the negative charge on phospholipids by attaching positively charged amino acid on the cytoplasmic side of inner membrane and then flipping the molecules to the outer leaflet of the inner membrane conferring resistance by electrostatic repulsion. Interestingly, we also found that a few nonmotile cells was enough to block the motility of motile cells, indicating an amplification of the inhibitory signal.;The next aim was to identify the underlying cause of swarm inhibition. To assess the behavioral outcome of mixed strains in swarm inhibition they were labeled with fluorescent tags or antibiotic markers. Surprisingly, we found that swarm inhibition was a result of killing of the motile cells by nonmotiles. Importantly, killing was Tra-dependent. omrA, a gene identified to be involved in the downstream response, conferred resistance to killing in the motile background, thus explaining the restoration of motility. On a closer examination of strain antagonism, motile cells were found to become filamentous with loss of DNA before lysis occurred. Interestingly, the killing phenotype correlated to sensitivity of the common wild-type strain, DK1622 or derived strains (motile strain), to their ancestral strains (nonmotile strain). For clarity, DK1622 is a reconstructed strain that has wild-type phenotypes for motility and development. The apparent reason behind sibling antagonism is a toxin/antitoxin system in a 200 kb region that contains prophage-like DNA element present in ancestral strains but missing in DK1622. Interestingly, the kill phenotype can be engineered toward non-siblings, including towards different species of myxobacteria. To do so, susceptible laboratory strains were programmed to kill environmental isolates by inserting a compatible traA allele from the environmental isolate. In summary, our results suggest that OME may play a role in modulating or 'policing' social interactions between cells. We hypothesize this function helps myxobacteria transition from individual cells into a coherent multicellular community of related bacteria.