Transcriptional changes when Myxococcus xanthus preys on Escherichia coli suggest myxobacterial predators are constitutively toxic but regulate their feeding

Abstract

Predation is a fundamental ecological process, but within most microbial ecosystems the molecular mechanisms of predation remain poorly understood. We investigated transcriptome changes associated with the predation of Escherichia coli by the myxobacterium Myxococcus xanthus using mRNA sequencing. Exposure to pre-killed prey significantly altered expression of 1319 predator genes. However, the transcriptional response to living prey was minimal, with only 12 genes being significantly up-regulated. The genes most induced by prey presence (kdpA and kdpB, members of the kdp regulon) were confirmed by reverse transcriptase quantitative PCR to be regulated by osmotic shock in M. xanthus, suggesting indirect sensing of prey. However, the prey showed extensive transcriptome changes when co-cultured with predator, with 40 % of its genes (1534) showing significant changes in expression. Bacteriolytic M. xanthus culture supernatant and secreted outer membrane vesicles (OMVs) also induced changes in expression of large numbers of prey genes (598 and 461, respectively). Five metabolic pathways were significantly enriched in prey genes up-regulated on exposure to OMVs, supernatant and/or predatory cells, including those for ribosome and lipopolysaccharide production, suggesting that the prey cell wall and protein production are primary targets of the predator's attack. Our data suggest a model of the myxobacterial predatome (genes and proteins associated with predation) in which the predator constitutively produces secretions which disable its prey whilst simultaneously generating a signal that prey is present. That signal then triggers a regulated feeding response in the predator.

Keywords: Myxobacteria; antimicrobial activity; mixed culture; outer membrane vesicles; predatome; transcriptome.

Fig. 1.

Gene expression under the six experimental conditions. (a) Experimental conditions for predator–prey RNAseq. M. xanthus predator (orange rods) and E. coli prey (white rods) were incubated separately or together, in either buffer (white background) or LBCY nutrient medium (yellow background). E. coli cells added to condition DEAD were heat-killed beforehand. (b) Gene expression (FPKM values) of the 7400 genes of the M. xanthus predator (left four columns) and the E. coli prey (right four columns), binned and coloured by magnitude, for each of the experimental conditions.

Fig. 2.

M. xanthus genes exhibiting differential expression (DE) on exposure to nutrients (Predator Nutrition), prey (LIVE) and pre-killed prey (DEAD), when compared with a nutrient-free control condition (Predator Starvation). DE gene numbers are shown for both high (a, b) and low (c, d) stringency filtering criteria [log(DE)>2 and DE>2, respectively]. Up-regulated genes (UR) are shown on the left (a, c) and down-regulated (DR) genes on the right (b, d).

Fig. 3.

Circos diagrams of the M. xanthus (a) and E. coli (b) genomes. Numbers around the diagrams each designate 10 000 bp. Black boxes represent genes encoded on the two DNA strands, red boxes designate genes encoding signalling and/or DNA-binding proteins, purple boxes are genomic islands, and grey boxes denote genes of secondary metabolite biosynthesis clusters. Nested rings represent genes differentially expressed (up-regulated in green and down-regulated in red) when comparing transcriptome datasets. The three comparisons in (a) are LIVE versus Predator Starvation (outermost), DEAD versus Predator Starvation, and Predator Nutrition versus Predator Starvation (innermost). The two comparisons in (b) are LIVE versus Prey Starvation and Prey Nutrition versus Prey Starvation.

Fig. 4.

E. coli genes exhibiting differential expression (DE) on exposure to M. xanthus cells (LIVE), OMVs and culture supernatant (when compared with the nutrient-free control condition Prey Starvation) with high (a, b) and low (c, d) stringency filtering criteria [log(DE)>2 and DE>2, respectively]. Up-regulated genes (UR) are shown on the left (a, c) and down-regulated (DR) genes on the right (b, d).

Fig. 5.

A model of transcriptional changes during predation of E. coli by M. xanthus. From left to right: prey cells close to the predator encounter its secretions such as OMVs (black circles). The predatory secretions attack the surface of the prey cell inducing changes in gene expression as the prey resist attack (1). Damage to the prey releases material (green circles) into the commons (2), which is sensed by the predator. The predator induces expression of genes for biomass assimilation, including hydrolases (yellow starbursts) which break down prey-derived material (3) for uptake by the predator (4). Pink arrows indicate constitutive secretion by the predator, green arrows denote the flow of prey-derived material, blue open arrows represent genes, while red arrows indicate the induction of gene expression and transport of gene products.