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Kin discrimination and outer membrane exchange in Myxococcus xanthus: Experimental analysis of a natural population.

ABSTRACT: In some species of myxobacteria, adjacent cells sufficiently similar at the adhesin protein TraA can exchange components of their outer membranes. The primary benefits of such outer membrane exchange (OME) in natural populations are unclear, but in some OME interactions, transferred OM content can include SitA toxins that kill OME participants lacking an appropriate immunity gene. Such OME-dependent toxin transfer across Myxococcus xanthus strains that differ only in their sitBAI toxin/antitoxin cassette can mediate inter-strain killing and generate colony-merger incompatibilities (CMIs)-inter-colony border phenotypes between distinct genotypes that differ from respective self-self colony interfaces. Here we ask whether OME-dependent toxin transfer is a common cause of prevalent CMIs and antagonisms between M. xanthus natural isolates identical at TraA. We disrupted traA in eleven isolates from a cm-scale soil population and assayed whether traA disruption eliminated or reduced CMIs between swarming colonies or antagonisms between strains in mixed cultures. Among 33 isolate pairs identical at traA that form clear CMIs, in no case did functional disruption of traA in one partner detectably alter CMI phenotypes. Further, traA disruption did not alleviate strong antagonisms observed during starvation-induced fruiting-body development in seven pairs of strains identical at traA. Collectively, our results suggest that most mechanisms of interference competition and inter-colony kin discrimination in natural populations of myxobacteria do not require OME. Finally, our experiments also indicate that several closely related laboratory reference strains kill some natural isolates by toxins delivered by a shared, OME-independent type VI secretion system (T6SS), suggesting that some antagonisms between sympatric natural isolates may also involve T6SS toxins.

SUBMITTER: Cossey SM

PROVIDER: S-EPMC6880969 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:UnlabelledMyxobacteria form complex social communities that elicit multicellular behaviors. One such behavior is kin recognition, in which cells identify siblings via their polymorphic TraA cell surface receptor, to transiently fuse outer membranes and exchange their contents. In addition, outer membrane exchange (OME) regulates behaviors, such as inhibition of wild-type Myxococcus xanthus (DK1622) from swarming. Here we monitored the fate of motile cells and surprisingly found they were killed by nonmotile siblings. The kill phenotype required OME (i.e., was TraA dependent). The genetic basis of killing was traced to ancestral strains used to construct DK1622. Specifically, the kill phenotype mapped to a large "polyploid prophage," Mx alpha. Sensitive strains contained a 200-kb deletion that removed two of three Mx alpha units. To explain these results, we suggest that Mx alpha expresses a toxin-antitoxin cassette that uses the OME machinery of M. xanthus to transfer a toxin that makes the population "addicted" to Mx alpha. Thus, siblings that lost Mx alpha units (no immunity) are killed by cells that harbor the element. To test this, an Mx alpha-harboring laboratory strain was engineered (by traA allele swap) to recognize a closely related species, Myxococcus fulvus. As a result, M. fulvus, which lacks Mx alpha, was killed. These TraA-mediated antagonisms provide an explanation for how kin recognition specificity might have evolved in myxobacteria. That is, recognition specificity is determined by polymorphisms in traA, which we hypothesize were selected for because OME with non-kin leads to lethal outcomes.ImportanceThe transition from single cell to multicellular life is considered a major evolutionary event. Myxobacteria have successfully made this transition. For example, in response to starvation, individual cells aggregate into multicellular fruiting bodies wherein cells differentiate into spores. To build fruits, cells need to recognize their siblings, and in part, this is mediated by the TraA cell surface receptor. Surprisingly, we report that TraA recognition can also involve sibling killing. We show that killing originates from a prophage-like element that has apparently hijacked the TraA system to deliver a toxin to kin. We hypothesize that this killing system has imposed selective pressures on kin recognition, which in turn has resulted in TraA polymorphisms and hence many different recognition groups.

Project description:Myxococcus xanthus cells coordinate cellular motility, biofilm formation, and development through the use of cell signaling pathways. In an effort to understand the mechanisms underlying these processes, the inner membrane (IM) and outer membrane (OM) of strain DK1622 were fractionated to examine protein localization. Membranes were enriched from spheroplasts of vegetative cells and then separated into three peaks on a three-step sucrose gradient. The high-density fraction corresponded to the putative IM, the medium-density fraction corresponded to a putative hybrid membrane (HM), and the low-density fraction corresponded to the putative OM. Each fraction was subjected to further separation on discontinuous sucrose gradients, which resulted in discrete protein peaks for each major fraction. The purity and origin of each peak were assessed by using succinate dehydrogenase (SDH) activity as the IM marker and reactivities to lipopolysaccharide core and O-antigen monoclonal antibodies as the OM markers. As previously reported, the OM markers localized to the low-density membrane fractions, while SDH localized to high-density fractions. Immunoblotting was used to localize important motility and signaling proteins within the protein peaks. CsgA, the C-signal-producing protein, and FibA, a fibril-associated protease, were localized in the IM (density, 1.17 to 1.24 g cm(-3)). Tgl and Cgl lipoproteins were localized in the OM, which contained areas of high buoyant density (1.21 to 1.24 g cm(-3)) and low buoyant density (1.169 to 1.171 g cm(-3)). FrzCD, a methyl-accepting chemotaxis protein, was predominantly located in the IM, although smaller amounts were found in the OM. The HM peaks showed twofold enrichment for the type IV pilin protein PilA, suggesting that this fraction contained cell poles. Two-dimensional polyacrylamide gel electrophoresis revealed the presence of proteins that were unique to the IM and OM. Characterization of proteins in an unusually low-density membrane peak (1.072 to 1.094 g cm(-3)) showed the presence of Ta-1 polyketide synthetase, which synthesizes the antibiotic myxovirescin A.

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Kin discrimination and outer membrane exchange in Myxococcus xanthus: Experimental analysis of a natural population.

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