Lactobacillus plantarum possesses the capability for wall teichoic acid backbone alditol switching

Abstract

BackgroundSpecific strains of Lactobacillus plantarum are marketed as health-promoting probiotics. The role and interplay of cell-wall compounds like wall- and lipo-teichoic acids (WTA and LTA) in bacterial physiology and probiotic-host interactions remain obscure. L. plantarum WCFS1 harbors the genetic potential to switch WTA backbone alditol, providing an opportunity to study the impact of WTA backbone modifications in an isogenic background.ResultsThrough genome mining and mutagenesis we constructed derivatives that synthesize alternative WTA variants. The mutants were shown to completely lack WTA, or produce WTA and LTA that lack D-Ala substitution, or ribitol-backbone WTA instead of the wild-type glycerol-containing backbone. DNA micro-array experiments established that the tarIJKL gene cluster is required for the biosynthesis of this alternative WTA backbone, and suggest ribose and arabinose are precursors thereof. Increased tarIJKL expression was not observed in any of our previously performed DNA microarray experiments, nor in qRT-PCR analyses of L. plantarum grown on various carbon sources, leaving the natural conditions leading to WTA backbone alditol switching, if any, to be identified. Human embryonic kidney NF-κB reporter cells expressing Toll like receptor (TLR)-2/6 were exposed to purified WTAs and/or the TA mutants, indicating that WTA is not directly involved in TLR-2/6 signaling, but attenuates this signaling in a backbone independent manner, likely by affecting the release and exposure of immunomodulatory compounds such as LTA. Moreover, human dendritic cells did not secrete any cytokines when purified WTAs were applied, whereas they secreted drastically decreased levels of the pro-inflammatory cytokines IL-12p70 and TNF-α after stimulation with the WTA mutants as compared to the wild-type.ConclusionsThe study presented here correlates structural differences in WTA to their functional characteristics, thereby providing important information aiding to improve our understanding of molecular host-microbe interactions and probiotic functionality.

Pathways and roles of wall teichoic acid glycosylation in Staphylococcus aureus

“…Several early studies already suggested that WTA GlcNAc may serve as an adsorption receptor for serogroup B phages such as Ф52A (Chatterjee, 1969;Park et al, 1974). However, S. aureus mutants lacking either ␣-or ␤-O-GlcNAc remained susceptible to most of the S. aureus phages while only the complete loss of WTA glycosylation resulted in phage resistance (Brown et al, 2012;Xia et al, 2010bXia et al, , 2011Xia and Wolz, 2013) (Table 1 and Fig. 1C).…”

“…Surprisingly, unlike transposon mutant K6, a targeted tarM deletion mutant (S. aureus RN4220 tarM) was still phage-susceptible and nuclear magnetic resonance (NMR) elucidation revealed the presence of ␤-O-GlcNAc substituted WTA in this strain further suggesting S. aureus encodes a second WTA GlcNAc transferase, potentially tagX or tarS (Brown et al, 2012). However, combined deletion of tarM and tagX did not alter phage susceptibility (Xia et al, unpublished data) and no in vitro WTA glycosyltransferase activity was found for TagX (Brown et al, 2012). When the other putative ␤-glycosyltransferase gene tarS of mutant K6 was sequenced it was found to be truncated because of a point mutation.…”

“…S. aureus NCTC8325-4 encodes ca. 20 putative glycosyltransferases and interestingly, two putative glycosyltransferases are co-localized within well studied WTA biosynthesis gene clusters namely tarS and tagX (Brown et al, 2012;Swoboda et al, 2010;Xia et al, 2010a) (Fig. 1B).…”

“…Based on the facts that mutant K6 that bears a transposon insertion in tarM lacked GlcNAc on WTAs and tarM complemented K6 mutant synthesizes ␣-O-GlcNAc on WTA, it was concluded that TarM is a WTA ␣-O-GlcNAc transferase (Xia et al, 2010b). Surprisingly, unlike transposon mutant K6, a targeted tarM deletion mutant (S. aureus RN4220 tarM) was still phage-susceptible and nuclear magnetic resonance (NMR) elucidation revealed the presence of ␤-O-GlcNAc substituted WTA in this strain further suggesting S. aureus encodes a second WTA GlcNAc transferase, potentially tagX or tarS (Brown et al, 2012). However, combined deletion of tarM and tagX did not alter phage susceptibility (Xia et al, unpublished data) and no in vitro WTA glycosyltransferase activity was found for TagX (Brown et al, 2012).…”

“…When the other putative ␤-glycosyltransferase gene tarS of mutant K6 was sequenced it was found to be truncated because of a point mutation. Moreover, TarS glycosylated RboP WTA polymers in vitro and deletion of both tarM and tarS resulted in complete absence of GlcNAc strongly suggesting tarS encodes the second S. aureus WTA glycosyltransferase (Brown et al, 2012).…”