Developmentally-Regulated Excision of the SPβ Prophage Reconstitutes a Gene Required for Spore Envelope Maturation in Bacillus subtilis

Abstract

Temperate phages infect bacteria by injecting their DNA into bacterial cells, where it becomes incorporated into the host genome as a prophage. In the genome of Bacillus subtilis 168, an active prophage, SPβ, is inserted into a polysaccharide synthesis gene, spsM. Here, we show that a rearrangement occurs during sporulation to reconstitute a functional composite spsM gene by precise excision of SPβ from the chromosome. SPβ excision requires a putative site-specific recombinase, SprA, and an accessory protein, SprB. A minimized SPβ, where all the SPβ genes were deleted, except sprA and sprB, retained the SPβ excision activity during sporulation, demonstrating that sprA and sprB are necessary and sufficient for the excision. While expression of sprA was observed during vegetative growth, sprB was induced during sporulation and upon mitomycin C treatment, which triggers the phage lytic cycle. We also demonstrated that overexpression of sprB (but not of sprA) resulted in SPβ prophage excision without triggering the lytic cycle. These results suggest that sprB is the factor that controls the timing of phage excision. Furthermore, we provide evidence that spsM is essential for the addition of polysaccharides to the spore envelope. The presence of polysaccharides on the spore surface renders the spore hydrophilic in water. This property may be beneficial in allowing spores to disperse in natural environments via water flow. A similar rearrangement occurs in Bacillus amyloliquefaciens FZB42, where a SPβ-like element is excised during sporulation to reconstitute a polysaccharide synthesis gene, suggesting that this type of gene rearrangement is common in spore-forming bacteria because it can be spread by phage infection.

De novo evolved interference competition promotes the spread of biofilm defectors

“…Normally, the excision of the SPβ prophage from the B. subtilis chromosome takes place before sporulation and allows reconstitution of the spsM gene involved in spore polysaccharide biosynthesis42. Sanchez-Vizuete et al 43.…”

The Exosporium Layer of Bacterial Spores: a Connection to the Environment and the Infected Host

“…These carbohydrates display structural similarity to those of the B. cereus family spores (49,50). Production and/or attachment of polysaccharides to the B. subtilis spore surface was shown to be spsM dependent (51). This determinant is the site of SP␤ prophage integration, and functional SpsM is produced only following excision of the prophage during sporulation (51).…”

“…Production and/or attachment of polysaccharides to the B. subtilis spore surface was shown to be spsM dependent (51). This determinant is the site of SP␤ prophage integration, and functional SpsM is produced only following excision of the prophage during sporulation (51). Spores from spsM- negative mutants had altered surface properties, including increased adherence to glass and enhanced aggregation in aqueous environments.…”

Genome-Wide Analysis of ResD, NsrR, and Fur Binding in Bacillus subtilis during Anaerobic Fermentative Growth by In Vivo Footprinting

“…Recent work showed that the site-specific recombination of the SP␤ region also occurs during sporulation, which generates the functional spore polysaccharide gene spsM from truncated yodU and ypqP (43). The sequences that ResD and NsrR bind cover the sites of recombination catalyzed by recombinases SprA (for SP␤) (43) and SpoIVCA (for sigK) (39) (Data Set S1). ResD and NsrR bind to the upstream site of ypqP that encodes the C-terminal part of SpsM (see Fig.…”