A Genetic Screen in Myxococcus xanthus Identifies Mutants That Uncouple Outer Membrane Exchange from a Downstream Cellular Response

Abstract

Upon physical contact with sibling cells, myxobacteria transiently fuse their outer membranes (OMs) and exchange OM proteins and lipids. From previous work, TraA and TraB were identified to be essential factors for OM exchange (OME) in donor and recipient cells. To define the genetic complexity of OME, we carried out a comprehensive forward genetic screen. The screen was based on the observation that Myxococcus xanthus nonmotile cells, by a Tra-dependent mechanism, block swarm expansion of motile cells when mixed. Thus, mutants defective in OME or a downstream responsive pathway were readily identified as escape flares from mixed inocula seeded on agar. This screen was surprisingly powerful, as we found >50 mutants defective in OME. Importantly, all of the mutations mapped to the traAB operon, suggesting that there may be few, if any, proteins besides TraA and TraB directly required for OME. We also found a second and phenotypically different class of mutants that exhibited wildtype OME but were defective in a responsive pathway. This pathway is postulated to control inner membrane homeostasis by covalently attaching amino acids to phospholipids. The identified proteins are homologous to the Staphylococcus aureus MprF protein, which is involved in membrane adaptation and antibiotic resistance. Interestingly, we also found that a small number of nonmotile cells were sufficient to block the swarming behavior of a large gliding-proficient population. This result suggests that an OME-derived signal could be amplified from a few nonmotile producers to act on many responder cells.

Sibling Rivalry in Myxococcus xanthus Is Mediated by Kin Recognition and a Polyploid Prophage

“…To track the fate of such cells, time-lapse microscopy was used 24 h after the cell mixture was plated. As previously reported (2,14), many of the cells at the swarm edge moved slowly or not at all (compare Movies S1 and S2 in the supplemental material), suggesting that those cells were sick or dead. In addition, in two cases, isolated cells lysed (see Movie S1).…”

“…We previously showed that swarm inhibition is Tra dependent (Fig. 1A) (2,14). That is, when either strain in the mixture contains a traA or traB mutation, swarm inhibition is abolished.…”

“…In contrast, the swarming and developmental behaviors of some motile strains can be antagonized by OME with some nonmotile strains (2). This antagonistic response is potent, as a ratio of 1 nonmotile cell to 50 motile cells blocks the latter from swarming (14). The nature of swarm inhibition is the focus of this study, in which we found that ancestral strains kill siblings that were derived from them.…”

Cell rejuvenation and social behaviors promoted by LPS exchange in myxobacteria

“…Although myxobacteria have a peculiar ability to produce long filamentous OM tubes, which some have argued are conduits for exchange, we find no evidence that these structures are required for OM exchange (23). Instead, we suggest that TraAB, the only known determinants for OME (50), catalyze the formation of prefusion junctions that lead to OM fusion and exchange.…”

Multifaceted Interfaces of Bacterial Competition

“…OME requires the production of an outer membrane protein complex, TraAB, in both the donor and recipient cell (126). TraAB appears to be the only component necessary to mediate OME (128) and, similarly to the BamA receptor in CDI systems, TraAB contains a polymorphic domain that limits OME to a narrow range of related targets (129). Given the functional similarities to CDI systems and the potential of OME to directly deliver toxic effectors into the envelope of target bacteria, it is not surprising that Myxobacteria use OME to mediate competition and engage in kin recognition.…”