Cellular Microbiology (2014) 16(7), 1014–1023

doi:10.1111/cmi.12304 First published online 2 June 2014

Microreview The biology of bacterial peptidoglycans and their impact on host immunity and physiology Richard Wheeler,1,2 Grégoire Chevalier,3 Gérard Eberl3* and Ivo Gomperts Boneca1,2** 1 Institut Pasteur, Biology and genetics of the bacterial cell wall Unit, Paris 75724, France. 2 INSERM, Avenir group, Paris 75015, France. 3 Institut Pasteur, Development of Lymphoid Tissues Unit, Paris 75724, France. Summary Peptidoglycans (PGN) are a constituent of the bacterial cell wall, and are shed as bacteria divide. The presence of PGN is therefore a marker of bacterial activity that has been exploited by both plants and animals to induce defence mechanisms. Pattern recognition receptors that recognize PGN are extremely well conserved throughout evolution and shown to play important and diverse role in the development, homeostasis and activation of the immune system. In addition, PGN can be detected beyond mucosal surfaces, and their receptor can be expressed in tissues and cells that are far from the niches where bacteria reside. Thus, PGN affects not only the host’s immunity, but also more generally the host’s physiology. In this review, we discuss the biochemistry and biology of PGN, and their intriguing effects on the development of the immune system and the host physiology.

receptor (PRRs) that recognize PGN, including NODlike receptor (NLRs), and PGN-recognition receptors (PGRPs) (Royet et al., 2011; Philpott et al., 2014). In both plants and animals, triggering of these receptors induces activation of the NF-κB pathway and consequently, an immune response. Much work has been accomplished on the activation and regulation of the immune system by PGN, partly as mutations in one NLR, Nod2, are associated with inflammatory bowel disease in humans (Lesage et al., 2002). The investigation of PGN is therefore of fundamental importance for understanding host–bacteria interactions and its clinical implications when this interaction is perturbed. Recognition of PGN by the host may determine not only immune parameters, but also physiological parameters such as metabolism and behaviour. PGN can be found in internal fluids and therefore can activate cells far from its source of production at mucosal surfaces (Clarke et al., 2010). Together with other microbe-associated molecular patterns (MAMPs), PGN may be part of the host’s normal set of regulatory molecules. In this review, we will bring together the current evidence supporting such a broad function of PGN, even though it is clear that much remains to be discovered. We will compile the existing knowledge on the biochemistry and biology of PGN in mice and humans, and suggest some avenues of exploration that should lead to an exciting new view of the host–microbe interaction, and its fundamental role in the development and homeostasis of the host.

Introduction The bacterial cell wall is a rigid polymer of sugar chains bound together by peptide bridges. This heteropolymer has to be at least partially degraded to allow for division, before it is reconstructed to yield mature daughter cells. In the process, small PGNs are released into the milieu, and constitute a marker for bacterial presence and activity. Metazoans have evolved a number of pattern recognition Received 15 March, 2014; revised 14 April, 2014; accepted 16 April, 2014. For correspondence. *E-mail [email protected]; Tel. (+33) 1 44 38 94 46; Fax (+33) 1 40 61 32 04; **E-mail bonecai@ pasteur.fr; Tel. (+33) 1 44 38 95 16; Fax (+33) 1 40 61 36 40. All authors contributed equally.

PGN structure The PGN network of the bacterial cell wall comprises glycan chains of β-1,4-linked N-acetylmuramic acid (MurNAc) and N-acetylglucosamine (GlcNAc), crosslinked by short peptide stems of L- and D-amino acids anchored to the lactyl moiety of MurNAc. The archetypical peptide structure is L-alanine, D-glutamate, a dibasic amino acid (typically meso-diaminopimelate (mDAP) in Gram-negative or L-lysine in Gram-positive bacteria), D-alanine and D-alanine. Peptide stems are typically cross-linked between the ε-amino group of the dibasic amino acid and the carboxyl group of D-alanine at the

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cellular microbiology

Bacterial peptidoglycans in the mammalian host fourth amino acid position, or more rarely between dibasic amino acids (Schleifer and Kandler, 1972). In Grampositive organisms, indirect cross-linking via an interpeptide bridge is common. During growth and maturation, the PGN is degraded by dedicated hydrolase enzymes causing PGN fragments (muropeptides) to be shed from the cell wall into the environment, a process termed PGN turnover. PGN hydrolases produced by the host and neighbouring microbiota can also liberate or modify PGN fragments. These fragments, released from the PGN layer, can stimulate host innate immune responses through Nod1 and Nod2 (Girardin et al., 2003a,b). In turn, bacterial PGN recycling pathways further modulate the bioavailability of soluble fragments, by removing them from the environment and thus preventing detection by the host (Johnson et al., 2013). The structural requirements of PGN recognition by Nod1 and Nod2 have been explored in detail (Girardin et al., 2003c; Wang et al., 2013) (Fig. 1). The presence of a glycan moiety is not required for Nod1 recognition. Human Nod1 (hNod1) is strongly activated by mesoDAPcontaining tripeptides (TriDAP), found predominantly in Gram-negative bacteria, whereas the minimal activating structure is the dipeptide γ-D-Glu-mesoDAP (iE-DAP). Murine Nod1 (mNod1) is activated by TetraDAP rather than TriDAP, although there is a degree of cross-over as hNod1 and mNod1 are weakly activated by TetraDAP and TriDAP respectively. An intact MurNAc moiety is an absolute requirement for Nod2 recognition, which is strongly activated by the muramyl dipeptides MurNAc-L-Ala-D-Glu (M-Di) and MurNAc-L-Ala-D-isoGln (MDP), present in

PGN of all Gram-positive and Gram-negative species (Girardin et al., 2003c). Nod2 also recognizes L-lysinecontaining muramyl tripeptide (MtriLys), found in the PGN of many Gram-positive bacteria. Administration of MtriLys in mice was protective against trinitrobenzene sulfonic acid-induced colitis, adding to evidence that Nod2 signalling can promote homeostasis and suppress inflammatory responses (Watanabe et al., 2004; Macho Fernandez et al., 2011). Nod2 was also stimulated by the tetrapeptide MTriLys-Asn in vitro, but the tetrapeptide had no protective effect in vivo. Other Nod-activating structures have been identified in vitro but have not been tested in vivo. These include the hNod1 ligand meso-lanthionine tripeptide, present in the PGN of the anaerobic bacterium Fusobacterium nucleatum, and the Nod2 ligand L-ornithine muramyl tripeptide, present in Gram-positive bacteria and some Gram-negative spirochaetes. Furthermore, the mesoDAP-type PGN of Gram-positive bacteria is typically modified by amidation of the mesoDAP residue ( DAPNH2 ). In vitro, MTriDAPNH2 weakly activates Nod2 (∼ 3.5-fold reduction compared to M-Di) and is not recognized by Nod1, although some Gram-positive PGNs weakly stimulate Nod1 in vitro due to low levels of nonamidated mesoDAP (Atrih et al., 1999; Girardin et al., 2003a). PGN modifications in host–bacteria interactions Lysozyme is a PGN muramidase and a cationic antimicrobial peptide present in the secretions of mammalian

Nod1

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MTriDAPNH2

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MTriLys

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L-Ala D-Glu

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D-Glu

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Fig. 1. The major agonists of human Nod1 and Nod2. The orange boxed area indicates the minimal structural motif recognized by Nod1. Orange ellipses indicate the major differences in the dibasic amino acid structure between muramyl tripeptides. MTriDAP, N-acetylmuramyl-L-alanyl-γ-D-glutamyl-meso-2,6-diaminopimelate; MTriDAPNH2 , MTriDAP with amidated meso-DAP; MtriLys, N-acetylmuramyl-L-alanyl-γ-D-glutamyl-L-lysine; MDP, N-acetylmuramyl-L-alanyl-γ-D-glutamate. © 2014 John Wiley & Sons Ltd, Cellular Microbiology, 16, 1014–1023

1016 R. Wheeler, G. Chevalier, G. Eberl and I. Gomperts Boneca mucosae, as well as in the phagolysosome and secretory granules of professional phagocytes. Lysozyme shapes the host immune response to bacteria by releasing Nodactivating PGNs and other MAMPs from the bacterial cell wall. However, bacteria can resist lysozyme activity by modification of the N-acetyl sugars of PGN. The major mechanism of lysozyme resistance is O-acetylation of the MurNAc C-6 hydroxyl group, found widely among bacterial pathogens (Moynihan and Clarke, 2010). In fact, O-acetylation of staphylococcal PGN is correlated with pathogenic potential (Bera et al., 2006), and Listeria monocytogenes O-acetylation contributes to survival in macrophages (Aubry et al., 2011). N-glycolylation or N-deacetylation have also been implicated in resistance to lysozyme activity (Raymond et al., 2005; Meyrand et al., 2007). Streptococcus pneumoniae combines MurNAc O-acetylation and GlcNAc deacetylation to resist lysis and death, although the PGN retains sufficient sensitivity for lysozyme-dependent release of the poreforming toxin pneumolysin and Nod2 ligands, suggesting a fitness cost, or a contribution of non-lethal PGN hydrolysis to virulence (Davis et al., 2011). GlcNAc deacetylation by L. monocytogenes also suppresses Nod1 activation, and in vitro deacetylated PGN of Helicobacter pylori is unable to induce Nod1 or Nod2 responses in HEK293 cells (Boneca et al., 2007). On the contrary, N-glycolylated muramic acid, found uniquely in actinomycetes such as Mycobacterium smegmatis and Mycobacterium tuberculosis, is a more potent activator of Nod2 as compared to MDP (Coulombe et al., 2009). PGN recognition proteins (PGRP)s are a family of bactericidal proteins carrying a conserved amidase domain, although, particularly in mammals, the amidase domain may lack enzymatic activity, functioning instead as a sensor of PGN. In drosophila, PGRP-LB and PGRP-SC amidases are scavenger proteins that negatively regulate inflammatory pathways by degrading biologically active muropeptides, such as those liberated by lysozyme (Paredes et al., 2011). Humans express four PGRPs of which only one member, PGLYRP-2, contains the amidase domain Zn2+-binding motif required for N-acetylmuramyl-L-alanine amidase activity. PGLYRP-2 is expressed in intraepithelial T lymphocytes and in the liver for secretion into the blood (Wang et al., 2003; Zhang et al., 2005; Duerr et al., 2011). The minimal ligand for PGLYRP-2 is muramyl tripeptide, thus PGLYRP-2 cannot function as a general scavenger of Nod2-ligands per se as it cannot hydrolyse MDP. However, PGLYRP-2 may regulate Nod1- versus Nod2-dependent responses by limiting the abundance of Nod2 tripeptide-ligands derived from Gram-positive bacteria, cleaving the essential glycan moiety required for Nod2 recognition, and thus favouring Nod1-induced responses by generating PGN peptides that bind Nod1 (Fig. 2). In support of such a function for

PGLYRP-2, pre-incubation of Staphylococcus aureus PGN with PGLYRP-2 reduces Nod2-stimulation in intestinal epithelial m-Iccl2 primary cell culture experiments (Duerr et al., 2011). On the other hand, the protective function of PGLYRP-2 against Salmonella enterica serovar Typhimurium infection in mice was Nod1/Nod2independent, suggesting that PGN processing by PGLYRP-2 is not involved in the response to Gramnegative pathogens (Lee et al., 2012). Other mammalian PGLYRPs are proposed to bind PGN at the division septum of Gram-positive bacteria and initiate lethal stress-responses through activation of two-component signal transductions systems (Kashyap et al., 2011). It is possible that in vivo, PGLYRP-2 amidase activity breaches the PGN barrier, facilitating an interaction with other PGLYRPs that contributes to host defence without bacterial lysis. PGN entry into cells Invasive intracellular pathogens release PGN directly into the cytosolic compartment, whereas extracellular pathogens deliver PGN to the cytosol through mechanisms such as the cagPAI-encoded type IV secretion system of H. pylori, outer membrane vesicles, or pore-forming toxins (Viala et al., 2004; Hruz et al., 2009; Kaparakis et al., 2010; Davis et al., 2011) (Fig. 2). However, the symbiotic microbiota does not normally contact the epithelial barrier directly. MAMPs diffuse through the mucous layer to reach the selectively permeable epithelial barrier. PGN fragments must then cross the epithelium and enter the cell cytosol via host-mediated uptake mechanisms in order to stimulate the cytosolic Nod-receptors. Selective transport mechanism could restrict PGN entry to the cytosol by steric-gating, which may explain why some Nod ligands defined in vitro have no apparent activity in vivo (Lee et al., 2009; Macho Fernandez et al., 2011). Three members of the SLC15 proton-oligopeptide co-transporter family, PepT1 (SLC15A1), PepT2 (SLC15A2) and PHT1 (SLC15A4), are involved in the transport of PGN di- and tripeptides, and muramyl peptides. Human (h)PepT1 is involved in the transport of MDP, and triDAP at high concentrations, when expressed ectopically in human intestinal epithelial cells (Vavricka et al., 2004; Ismair et al., 2006; Dalmasso et al., 2010), while PepT2 was implicated in NOD1activation by γ-iE-DAP transport in human lung epithelial cells (Swaan et al., 2008). PHT1 expression has been implicated in the cytosolic entry of TriDAP internalized through clathrin-mediated endocytosis (Marina-Garcia et al., 2008; Lee et al., 2009). Transmembrane channels may also function as a general mechanism for PGN to enter the cytosol. Pannexin-1 activity facilitates cytosolic entry of endocytosed MDP-rhodamine, although it is © 2014 John Wiley & Sons Ltd, Cellular Microbiology, 16, 1014–1023

Bacterial peptidoglycans in the mammalian host

Bacterial Host hydrolases hydrolases Lysosyme, PGLYRP-2(?)

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Transport e.g. SLC15 family

NOD activation NOD1

Phagocytic lysis

Pore-forming toxins e.g. pneumolysin

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Cytosol

Fig. 2. Proposed pathways of muropeptide entry into the cytosol and Nod-activation. PGN fragments can be delivered to the cytosol through specific virulence mechanisms, e.g. type-IV secretion systems, pore-forming toxins or outer membrane vesicles (OMVs). In the extracellular milieu, PGN fragments released by cell wall turnover, by hydrolases produced by the host and competing flora, or due to bacterial lysis, may be endocytosed and released to the cytosol via active transport or transmembrane channels. Phagocytosed bacteria may further produce muropeptides in the phagosome and following lysis in phagolysosomes. Invasive bacteria activate Nod receptors directly by shedding muropeptides into the cytosol.

© 2014 John Wiley & Sons Ltd, Cellular Microbiology, 16, 1014–1023

1018 R. Wheeler, G. Chevalier, G. Eberl and I. Gomperts Boneca unclear whether pannexin-1 acts as an MDP transporter or induces endosomal changes that triggers MDP release (Marina-Garcia et al., 2008). As most PGN transport data have been inferred in transfected cell lines, there is a pressing need to explore PGN transport mechanisms in Nod-expressing cell lines that have been unequivocally demonstrated to naturally internalize PGN fragments. One such cell line (Hec1B) has been recently identified (Zarantonelli et al., 2013). Furthermore, it remains to be demonstrated that these putative PGN transporters play a role in PGN internalization in vivo. PGN dissemination to distal tissues Whereas it was shown that PGN fragments are present in CSF and urine of patients with sleeping disorders, the presence of PGN fragments in host tissues and serum in the absence of invasive disease has been a matter of debate (Fencl et al., 1971; Krueger et al., 1984; Johannsen et al., 1989). Using muramic acid as a marker for the presence of PGN, fragments were reported in the brain, liver and kidney of rats by thin-layer chromatography, and in human spleen by reverse-phase HPLC, initiating speculation that PGN originating from the gut microbiota can diffuse systemically (Sen and Karnovsky, 1984; Hoijer et al., 1995). However, the specificity of these detection methods has been criticized, and alternative approaches combining gas-chromatography and mass spectrometry failed to detect PGN in rabbit serum, or rat brain and spleen (Fox and Fox, 1991; Kozar et al., 2000). Furthermore, radiolabelled MDP was rapidly cleared from the host when injected intravenously, and only trace amounts persisted in most host tissues (Parant et al., 1979). More recent studies combining chromatography and NMR have detected PGN in human plasma and bovine serum (Xu et al., 2008). Furthermore, Clarke et al. (2010) demonstrated that [3H]mesoDAP-labelled Escherichia coli PGN, when delivered intragastrically, translocates across the gut epithelium and is detectable in the sera and bone marrow. Critically, these PGN fragments are bioactive, as sera from mice induced Noddependent NF-κB activation in HEK293 reporter assays (Clarke et al., 2010). Recent data strongly suggests that PGN disseminates systemically from the gut mucosa and has the potential to affect distal tissue homeostasis and immunity. This hypothesis is also supported by the broad distribution of Nod1 and Nod2 receptors. Nod1 is expressed in most adult tissues (Bertin et al., 1999; Inohara et al., 1999; Rosenzweig et al., 2008; Scott et al., 2010), while Nod2 is expressed in immune cells, where it predominates in peripheral blood monocytes, granulocytes and dendritic cells, as well as in astrocytes of the brain and in paneth cells of the gut (Ogura et al., 2001; Sterka et al., 2006).

Importantly, Nod1 and Nod2 expression overlaps with that of the putative peptidoglycan transporters PepT1, PepT2 and PHT1. The impact of PGN on development of the immune system Nod1 and Nod2 monitor the cytoplasm for the presence of PGN. Activated NLRs then initiate an inflammatory cytokine response through the signalling molecule RIP2 that leads to resistance to infection. They also initiate the autophagy system that leads to clearance of intracellular bacteria (Philpott et al., 2014). In mice deficient for Nod1, Nod2, or RIP2, defects have been found in the production of antimicrobial peptides, paracellular permeability of epithelial cells, and chemokine secretion, defects that may account for the impact of these mutations on the response to infection and on IBD. However, an unexpected consequence of defective antimicrobial peptide production is the decreased formation of lymphoid follicles in the intestinal lamina propria (Bouskra et al., 2008). As Philpott and colleagues have recently written a comprehensive review of the effect of PGN on immune responses (Philpott et al., 2014), we will focus our review on the impact of PGN on the development and structure of the immune system. In the fetus, development of lymph nodes and Peyer’s patches is initiated by the recruitment of lymphoid tissue inducer (LTi) cells to sites pre-determined by specialized stromal cells (Mebius, 2003). LTi cells form clusters that activate stromal cells and induce the recruitment of lymphocytes and dendritic cells (DCs) to form mature lymphoid tissues. After birth, similar clusters of LTi cells, termed cryptopatches, form in the intestinal lamina propria (Kanamori et al., 1996; Eberl and Littman, 2004). Bacteria colonizing the intestine activate cryptopatches to recruit B cells and form isolated lymphoid follicles (ILFs) (Hamada et al., 2002), which manufacture IgA against the symbiotic microbiota, thus establishing a negative feedback loop (Tsuji et al., 2008). In germfree mice, cryptopatches develop but not ILFs, while in mice with microbiota, the number of intestinal bacteria increases 10-fold in the absence of cryptopatches and ILFs (Bouskra et al., 2008). We found that LTi cells in cryptopatches, and B cells, are activated through the chemokine receptor CCR6 by β3-defensin or CCL20 expressed by epithelial cells. This expression is in turn induced by PGN recognized by Nod1, but not Nod2. As a consequence, mice that do not express Nod1 in epithelial cells show a significant decrease in the formation of ILFs as well as perturbed intestinal homeostasis (Bouskra et al., 2008). A similar phenomenon was reported in the Hawaiian squid Euprymna scolopes in its symbiotic relationship with the luminescent bacterium Vibrio fischeri (Koropatnick © 2014 John Wiley & Sons Ltd, Cellular Microbiology, 16, 1014–1023

Bacterial peptidoglycans in the mammalian host et al., 2004; McFall-Ngai et al., 2009). The bacterial symbiont penetrates ventral crypts and releases PGN and lipid A, which together induce apoptosis of superficial epithelial cells and infiltration of macrophage-like haemocytes. As superficial epithelial cells mediate the recruitment of V. fischeri (Kremer et al., 2013), their PGNinduced death shuts off further colonization by potential competitors (Koropatnick et al., 2014). Interestingly, PGN by itself can induce remodelling of the crypts, demonstrating that, similar to the mouse lymphoid tissue system, the squid system integrates PGN in the development of its host–microbe symbiosis. A report showing that PGN enhances the activity of neutrophils in bone marrow suggests that PGN may also have an effect on haematopoiesis. Neutrophils from NOD1-deficient mice or antibiotic-treated mice killed bacteria less efficiently, while full effector activity was induced by PGN provided to wild-type mice (Clarke et al., 2010). As PGN derived from radiolabelled intestinal bacteria was detected in serum and bone marrow, it is possible that PGN directly modulates the activity of neutrophils in the bone marrow where they are generated. An effect of PGN on the development of myeloid cells remains however to be formally assessed. Recently, it was shown that intestinal microbiota contributes to myelopoiesis and, as a consequence, to resistance to systemic infection by L. monocytogenes (Khosravi et al., 2014). Furthermore, intestinal inflammation induces IL-23-dependent bone marrow myelopoiesis at the expense of erythro- and lymphopoiesis (Griseri et al., 2012). As intestinal inflammation increases intestinal permeability, it is possible that serum concentrations of PGN are increased in this context and regulate haematopoiesis. In the same vein, another MAMP and ligand of TLR9, CpG, affects pro-B cell expansion in the bone marrow (Lalanne et al., 2010). These data show that MAMPs produced by the symbiotic microbiota, including PGN, can affect the development and activation of immune cells in the bone marrow and thereby modulate systemic immunity. The impact of PGN on host physiology Systems other than the immune system are affected by PGN, even though leucocytes may be primarily responding to PGN and relaying signals (Fig. 3). MDP synergistically enhances osteoclast formation induced by IL-1α and TNFα through Nod2-dependent RANKL expression in osteoblasts (Yang et al., 2005). This effect of PGN was necessary but not sufficient, as MDP alone could not induce osteoclast formation. Furthermore, MDP induces fever in guinea pigs (Roth et al., 1997), a physiological defence mechanism against infection. TNFα and IL-6 are the likely host inducers of fever induced by MDP, as high serum levels of these cytokines were detected soon after © 2014 John Wiley & Sons Ltd, Cellular Microbiology, 16, 1014–1023

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Nod1/2-expressing cells in brain, bones?

In synergy with

TNF-α, IL-6, IL-1

Fever

Sleep

Osteogenesis

Fig. 3. Impact of PGN beyond the immune system. PGN produced by symbiotic microbiota may ‘leak’ into the bloodstream and reach organs distant to the gut, such as the bones and the brain. There, the effects of PGN on osteogenesis, induction of fever and sleep are mediated by pro-inflammatory immune mediators, such as TNFα, IL-6 and IL-1, which may be produced by cells from the immune system present in these tissues.

MDP injection and prior to temperature increase. However, the vagal nerve that innervates mainly the heart, liver, pancreas and intestine, could also relay signals to induce fever since MDP-induced fever is partially attenuated in vagotomized guinea pigs (Goldbach et al., 1997). Another intriguing effect of PGN was reported on sleeping patterns. The somnogenic properties of PGN were first discovered in experiments to isolate sleep-promoting substances from sleep-deprived brains and urine from sleepdeprived patients. As prolonged wakefulness results in sleepiness, it was hypothesized that this state would accumulate somnogenic factors. This hypothesis was formulated more than a century ago (Kubota, 1989), and the so-called Factor S eventually purified and described as a muramyl peptide (MDP) in the 1980s (Krueger et al., 1982). MDP showed a powerful somnogenic effect, as 1 pmol of MDP injected into the brain ventricles could enhance slow-wave sleep in rabbits for 6 to 12 h (Krueger et al., 1984). It was also shown that intravenous injection of isolated MDP elicits sleep responses similar to those observed with viable bacteria (Johannsen et al., 1990) and MDP derived from bacteria digested by macrophages

1020 R. Wheeler, G. Chevalier, G. Eberl and I. Gomperts Boneca (Johannsen et al., 1991). Interestingly, sleep deprivation in rats leads to bacterial translocation from the intestine to the mesenteric lymph nodes (Everson and Toth, 2000), supporting the possibility that the gut microbiota is a source of systemic MDP that may reach the brain. It is likely that sleep responses induced by MDP are mediated by IL-1β and TNF-α as pre-treatments with inhibitors of those cytokines attenuate MDP-induced sleep (Imeri et al., 1993; Takahashi et al., 1996), and both IL-1β and TNF-α enhance sleep, indicating again that the nonimmune effects of MDP may be mediated by immune effectors. Conclusion The gut microbiota contains almost 1000 different bacterial species producing a diverse peptidoglycome. Using cutting-edge analytical tools, such as liquidchromatography coupled to mass spectrometry, it is now possible to establish the peptidoglycome of an individual in the intestine, serum and organs. Given the effects of PGN on immunity and physiology, discovered and yet to be discovered, determination of the peptidoglycome will be predictive of particular effects of the symbiotic microbiota on its host, and a consequence of the mucosal permeability, as well as of the composition and activity of the microbiota. We therefore propose ‘peptidoglycomics’ to be developed as a valuable diagnostic tool of the state of the host–microbiota interaction. Furthermore, the activation and inhibition of Nod1 and Nod2 through agonists and antagonists may be the basis for the development of therapies to pathologies involving these PRRs, such as IBD. In support of this view, Watanabe (Watanabe et al., 2004), Macho Fernandez (Macho Fernandez et al., 2011) and co-workers have independently demonstrated that PGN signalling through Nod2 can suppress the pathological inflammatory response. Modulation of Nod1 and Nod2 may also be the basis for therapies against physiological abnormalities such as sleeping disorders, even though the mechanisms involved still need to be uncovered. Acknowledgements We wish to thank all our colleagues from our two laboratories for support and fruitful discussions. This work has been supported by grants from the Agence Nationale de la Recherche, the Fond de la Recherche Médicale, the Institut Pasteur and Danone Research. Furthermore, the IGB laboratory is supported by an ERC starting grant (PGNformSHAPEtoVIR n°202283). We declare no conflicts of interests.

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The biology of bacterial peptidoglycans and their impact on host immunity and physiology.

Peptidoglycans (PGN) are a constituent of the bacterial cell wall, and are shed as bacteria divide. The presence of PGN is therefore a marker of bacte...
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