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International Journal of Medical Microbiology 300 (2010) 148154

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International Journal of Medical Microbiology

journal homepage: www.elsevier.de/ijmm

Mini Review

The wall teichoic acid and lipoteichoic acid polymers of Staphylococcus aureus
Guoqing Xia, Thomas Kohler, Andreas Peschel n
Division of Cellular and Molecular Microbiology, Institute of Medical Microbiology and Hygiene, University of T ubingen, Elfriede-Aulhorn-Strae 6, D-72076 T ubingen, Germany

a r t i c l e in f o

a b s t r a c t
Staphylococci and most other Gram-positive bacteria incorporate complex teichoic acid (TA) polymers into their cell envelopes. Several crucial roles in Staphylococcus aureus tness and cell wall maintenance have been assigned to these polymers, which are either covalently linked to peptidoglycan (wall teichoic acid, WTA) or to the cytoplasmic membrane (lipoteichoic acid, LTA). However, the exact TA structures, functions, and biosynthetic pathways are only supercially understood. Recently, most of the enzymes mediating TA biosynthesis have been identied and mutants lacking or with dened changes in WTA or LTA have become available. Their characterization has revealed crucial roles of TAs in protection against harmful molecules and environmental stresses; in control of enzymes directing cell division or morphogenesis and of cation homeostasis; and in interaction with host or bacteriophage receptors and biomaterials. Accordingly, several in vivo studies have demonstrated the importance of WTA and LTA in S. aureus colonization, infection, and immune evasion. TAs and enzymes required for TA biosynthesis represent attractive candidates for novel vaccines and antibiotics and are targeted by recently developed antibacterial therapeutics. & 2009 Elsevier GmbH. All rights reserved.

Keywords: Wall teichoic acid Lipoteichoic acid Glycopolymers Cell wall Microbehost interaction Gram-positive bacteria Staphylococci

Introduction Staphylococcus aureus is extremely successful in colonizing and infecting human and animal hosts. This capacity depends on adaptation to several, completely different habitats such as (i) human skin involving exposure to dryness, high salt concentration, and antimicrobial fatty acids; (ii) human nares with moist surfaces and high amounts of antimicrobial peptides; and, (iii) during infection, otherwise sterile host tissues containing phagocytes and the entire arsenal of antimicrobial host defenses et al., 2007). Accordingly, S. aureus cells have to (Lowy, 1998; Gotz protect themselves effectively against many different harmful molecules and environmental stress. This task is particularly challenging for the cell envelope, which is directly accessible to small molecules. Gram-positive bacteria have been shown to modify their cell envelopes in multiple ways to prevent the access and damage by antimicrobial molecules. In addition to covalent modication of peptidoglycan (PG) and phospholipids, the production of protective capsule and slime polymers, and the secretion of proteinaceous evasins (Foster, 2005; Kraus and Peschel, 2008), most Gram-positive bacteria have teichoic acids (TAs) or related glycopolymers that play crucial roles in bacterial survival under disadvantageous conditions and in other basic cellular processes (Weidenmaier and Peschel, 2008; Kohler et al.,

Corresponding author. Tel.: + 49 7071 298 1515; fax: + 49 7071 293 435. E-mail address: andreas.peschel@uni-tuebingen.de (A. Peschel).

2009b). TAs are usually constitutively produced and either connected to PG (wall teichoic acids, WTA) or to the cytoplasmic membrane (lipoteichoic acids, LTA). S. aureus and most other Gram-positive bacteria produce both TA types (Fig. 1A). Several recent studies have shed new light on the functions and biosynthesis of WTA and LTA, which are summarized and discussed in this overview article. Most TAs exhibit zwitterionic properties because of the presence of negatively charged phosphate groups and additional D-alanine residues on the repeating units, which have free positively charged amino groups (Neuhaus and Baddiley, 2003; Weidenmaier and Peschel, 2008; Kohler et al., 2009b). The known WTA structures vary widely between bacterial species and often even between clonal groups. S. aureus TAs are composed of repetitive polyol phosphate subunits such as ribitol phosphate (Rbo-P) or glycerol phosphate (Gro-P). As shown in Fig. 1B, S. aureus WTA is covalently linked to the 6-OH of N-acetyl muramic acid (MurNAc) via a disaccharide composed of Nacetylglucosamine (GlcNAc)-1-P and N-acetylmannosamine (ManNAc), which is followed by two units of Gro-P (Araki and Ito, 1989; Brown et al., 2008). The actual WTA polymer is composed of 1140 Rbo-P repeating units in most S. aureus strains, e.g. S. aureus Copenhagen (Sanderson et al., 1962) or of Gro-P repeating units, e.g. in S. aureus strain 187 (Endl et al., 1983). Strain NM8m, a potent biolm producer, appears to produce even both, poly-Rbo-P and poly-Gro-P WTA (Vinogradov et al., 2006). Staphylococcus epidermidis and some other staphylococcal species produce simpler WTA structures with Gro-P units forming the

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entire polymer (Sadovskaya et al., 2004; Endl et al., 1983). Very complex, hexose phosphate-containing WTAs are found, e.g. in Staphylococcus hyicus or Staphylococcus auricularis (Endl et al., 1983).

LTA polymers are attached to the cytoplasmic membrane via a glycolipid anchor, which is a diglucosyl diacylglycerol in S. aureus and most other staphylococcal species (Wicken and Knox, 1975; Fischer, 1988). The most often found LTA backbone is formed by Gro-P repeating units. As a likely consequence of the unique biosynthetic pathway, the structures of LTA are usually less diverse than those of WTA (Fischer et al., 1994 ). At the 2-hydroxyl group of the glycerol, S. aureus LTA is substituted with D-alanyl ester or a-GlcNAc (Fischer, 1988).

Why does S. aureus produce TAs? The actual functions of TAs and the reasons why most Grampositive bacteria produce both LTA and WTA at the same time remain incompletely understood. WTA has been shown to be dispensable for viability of S. aureus and Bacillus subtilis under laboratory conditions (Weidenmaier et al., 2004; DElia et al., 2006) but to play important roles during colonization and infection in vivo (Weidenmaier et al., 2004, 2005; Dubail et al., 2006; Bizzini et al., 2007; DElia et al., 2009). In contrast, LTA is dispensable only at temperatures below 30 1C, and it turned out to be impossible to delete both WTA and LTA at the same time since the two types of TA appear to compensate for one another to some extent (Oku et al., 2009; Schirner et al., 2009). The known TA functions can be classied in three major themes: (i) protection against harmful molecules and environmental stresses, (ii) control of enzyme activities and cation concentrations in the cell envelope, and (iii) binding to receptors and surfaces (Table 1). While WTA, LTA or related polymers are known to bind cell wall proteins, S-layers, or mycolic acids in other Gram-positive bacteria, staphylococci do not seem to employ LTA and WTA to tether additional protection layers. Nevertheless, WTA and LTA are crucial for preventing the passage of harmful molecules through the PG layers in S. aureus. WTA contributes to lysozyme resistance in concert with MurNAc O-acetylation probably by preventing binding of the enzyme to the glycan strands of PG (Bera et al., 2007). The highly hydrophilic WTA provides resistance against highly hydrophobic antimicrobial fatty acids from human skin by reducing the bacterial afnity for fatty acids and may thus enable S. aureus and other skin-colonizing Gram-positive bacteria to survive on human skin. Of note, the level of resistance mediated

Fig. 1. Schematic localization in the cell envelope (A) and structure (B) of S. aureus wall teichoic acid (WTA) and lipoteichoic acid (LTA). P, phosphate; D-Ala, D-alanine; GlcNAc, N-acetylglucosamine; ManNAc, N-acetylmannosamine; MurNAc, N-acetylmuramic acid; Glc, Glucose.

Table 1 Known and proposed functions of S. aureus WTA and LTA. Function Protection against cell damage Resistance to antimicrobial peptides (CAMPs) Resistance Resistance Resistance Resistance Resistance to to to to to cationic antibiotics (e.g. vancomycin) lysozyme antimicrobial fatty acids heat stress low osmolarity TA References

D-ala of TA D-ala of TA WTA WTA WTA, LTA LTA LTA WTA, LTA WTA LTA LTA LTA WTA LTA, WTA

Peschel et al., 1999; Collins et al., 2002; Koprivnjak et al., 2002; Weidenmaier et al., 2005 Peschel et al., 2000 Bera et al., 2007 Kohler et al., 2009a Hoover and Gray, 1977; Vergara-Irigaray et al., 2008; Oku et al., 2009 Oku et al., 2009 Grundling and Schneewind, 2007b; Oku et al., 2009 Bierbaum and Sahl, 1985; Fedtke et al., 2007; Vergara-Irigaray et al., 2008 Weidenmaier et al., 2004, 2005, 2008 Greenberg et al., 1996; Dunne et al., 1994 Polotsky et al., 1996; Lynch et al., 2004; Nahid and Sugii, 2006 Morath et al., 2001, 2002; Hermann et al., 2002; Hoebe et al., 2005; Draing et al., 2008 Chatterjee, 1969; Park et al., 1974 Gross et al., 2001; Fedtke et al., 2007; Vergara-Irigaray et al., 2008

Controlling protein machineries in the cell envelope Cell division site placement Autolysin activity Mediating interaction with receptors and biomaterials Adherence to epithelial and endothelial cells Binding to scavenger receptors (e.g. SrA) Activation of the complement system via MBL and colin Induction of inammation via TLR2, CD36, CD14, LBP Serving as phage receptor Mediating attachment to biomaterials and biolm formation

CAMP, cationic antimicrobial peptide; SrA, scavenger receptor A; MBL, mannose-binding lectin; TLR2, toll-like receptor 2; LBP, lipopolysaccharide-binding protein.

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by WTA correlated with the length and hydrophobicity of the antimicrobial fatty acids (Kohler et al., 2009a). The net charge of WTA and LTA governs the susceptibility of S. aureus and many other Gram-positive bacteria to cationic antimicrobial peptides (CAMPs) such as defensins, cathelicidins, and kinocidins (Peschel et al., 1999; Collins et al., 2002; Koprivnjak et al., 2002; Weidenmaier et al., 2005) and to cationic antibiotics such as vancomycin (Peschel et al., 2000). In order to reduce the highly negative net charge in the cell envelope, WTA and LTA are modied with D-alanine, and mutants lacking this modication are highly susceptible to CAMPs (Peschel et al., 1999). LTA- or WTA-decient S. aureus mutants are sensitive to high temperatures (Hoover and Gray, 1977; Vergara-Irigaray et al., 2008; Oku et al., 2009), and LTA is important for S. aureus survival under lowosmolarity conditions for currently unclear reasons (Oku et al., 2009). Interestingly, certain antimicrobial host proteins seem to make use of WTA, since WTA-decient S. aureus mutants are more resistant to mammalian group IIA phospholipase A2 and beta defensin 3 than wild-type strains (Koprivnjak et al., 2008). TAs seem to affect the function of important housekeeping enzyme machineries in Gram-positive cell envelopes. It is assumed that LTA interacts with components of the membranebound cell division machinery and contributes to its proper positioning or regulation because depletion of LTA leads to S. aureus cells with distorted shapes and division sites (Grundling and Schneewind, 2007a; Oku et al., 2009). In accord with this nding, the LTA polymerase LtaS has been shown to be mainly located in the division sites in both vegetative and sporulating B. subtilis cells (Schirner et al., 2009). WTA and LTA also play a profound role in controlling the activity of autolysins (Rice and Bayles, 2008), which show high afnities for LTA in vitro (Bierbaum and Sahl, 1985, 1987; Giudicelli and Tomasz, 1984). S. aureus mutants with reduced LTA amount or lacking LTA exhibited strongly reduced autolysis (Fedtke et al., 2007; Oku et al., 2009), and mutants lacking WTA have profoundly increased autolysis activities compared to wild-type cells (Vergara-Irigaray et al., 2008), indicating a complex regulatory interaction of LTA, WTA, and autolysins. Accordingly, WTA inhibits the side wall localization of the LysM domain containing autolysin LytF, the DLendopeptidase activity of which is required for cell separation in B. subtilis (Yamamoto et al., 2008). The capacity of TAs to control cell envelope enzymes may be related to the ion exchanger-like properties of TAs and the capacity to bind magnesium ions with particularly high afnity (Heptinstall et al., 1970). Accordingly, an LTA-depleted B. subtilis mutant shows a higher requirement for Mg2 + and increased susceptibility to toxic Mn2 + (Schirner et al., 2009). TAs are exposed at the bacterial surface and can interact with host cells, bacteriophages, or inert surfaces (Table 1). A strong impact on adherence to biomaterials and biolm formation has been observed in S. aureus and Enterococcus faecalis mutants decient in TA alanyl esters or with altered TA amounts in in vitro and in vivo models (Gross et al., 2001; Fabretti et al., 2006; Fedtke et al., 2007; Vergara-Irigaray et al., 2008). Many staphylococcal bacteriophages use WTA as receptors probably by lectin-mediated binding to the GlcNAc residues on WTA (Chatterjee, 1969; Park et al., 1974). As a consequence, differences in WTA structure play a crucial role in staphylococcal phage typing (Pantucek et al., 2004). WTA and LTA have also been implicated in S. aureus interaction with a wide range of host receptors and are thought to shape the entire colonization and infection process, ranging from initial adherence, activation of innate immunity, to the elicitation of adaptive immune reactions (Weidenmaier and Peschel, 2008). WTA contributes to binding of S. aureus to epithelial and endothelial cells most probably via lectin-like receptors, which remain to be identied (Weidenmaier et al., 2004, 2005, 2008).

Accordingly, S. aureus mutants lacking WTA have lost the ability to colonize the nose in animal models or to leave the bloodstream and infect sub-endothelial tissues in endovascular infections. LTA has been shown to bind to soluble C-type lectins such as the mannose-binding lectin (MBL) and L-colin, thereby activating the lectin-initiated complement pathway, which leads to bacterial opsonization and release of chemotactic complement split products (Polotsky et al., 1996; Lynch et al., 2004; Nahid and Sugii, 2006). A number of studies have indicated that LTA activates the innate immune system via Toll-like receptor 2 (TLR2) (Morath et al., 2001, 2002; Hermann et al., 2002; Hoebe et al., 2005; Tapping and Tobias, 2003; von Aulock et al., 2007; Draing et al., 2008). However, most of the commonly used LTA preparations are contaminated with bacterial lipopeptides, which account for a large percentage of the TLR2-mediated proinammatory activity (Hashimoto et al., 2006, 2007; Zahringer et al., 2008), and the proinammatory potency of LTA remains a matter of vigorous debate. S. aureus TAs are well-known targets for antibodies (Kumar et al., 2005), and TAs have been developed as vaccines (Schaffer and Lee, 2009). Indeed, promising results have been obtained through passive vaccination with a humanized monoclonal antibody targeting staphylococcal LTA (Weisman, 2007; Weisman et al., 2009). Previous studies have also led to the assumption that the zwitterionic S. aureus WTA might be processed and presented by antigen-presenting cells and serve as an antigen for specic T cells (Tzianabos et al., 2001). Thus, TAs are most critical in S. aureushost interaction and represent attractive targets for new anti-colonization or anti-infection strategies.

How does S. aureus synthesize WTA and LTA? Despite of the structural similarity, the biosynthesis of WTA and LTA relies on profoundly different pathways and precursor molecules. S. aureus requires at least 12 genes for biosynthesis of poly-Rbo-P WTA while only three genes seem to be required for biosynthesis of poly-Gro-P LTA backbones. Further genes are required for subsequent modication with D-alanine and hexoses. In contrast, synthesis of the extremely complex WTA-like polymer of Streptococcus agalactiae representing the Lanceeld group B antigen is supposed to depend on more than 120 genes (Sutcliffe et al., 2008). Again, it remains enigmatic why Gram-positive bacteria invest so much genetic information and energy in the maintenance of different pathways ultimately leading to very similar polymers. The biosynthetic pathways of WTA and related PG-anchored polymers share some widely conserved principles such as the use of undecaprenylphosphate (C55-P) as lipid carrier during the assembly process and the genomic organization of most biosynthetic genes in clusters (Brown et al., 2008; Xia and Peschel, 2008; Meredith et al., 2008). Biosynthesis of WTA is initiated on C55-P, which is also used for the biosynthesis of PG or capsular polysaccharide at the inner leaet of the cytoplasmic membrane. Fig. 2A shows the current model of S. aureus WTA biosynthesis, which can be divided into ve steps. The nomenclature of genes and proteins has been subject to some confusion recently. We tried to omit renaming here and use names that follow the function and allocation of genes in gene clusters. Accordingly, the gene acronym tag is used for genes involved in conserved steps that are shared by both, poly-Gro-P and poly-Rbo-P WTA biosynthetic pathways while tar is used only for those genes that are additionally required for incorporation of Rbo-P units (Xia and Peschel, 2008). WTA biosynthesis is initiated by the synthesis of a canonical disaccharide linkage unit, which requires the enzymes TagO and TagA transferring GlcNAc-1-phosphate and

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Fig. 2. Pathways of S. aureus wall teichoic acid (WTA) biosynthesis (A), lipoteichoic acid (LTA) biosynthesis (B), and D-alanine incorporation into LTA and WTA (C). Reproduced from Kohler et al. (2009b) with permission. CDP-Gro, cytidyldiphosphate-glycerol; CDP-Rbo, cytidyldiphosphate-ribitol; Glc, glucose; GlcNAc, Nacetylglucosamine; Gro, glycerol; Gro-P, glycerolphosphate; ManNAc, N-acetylmannosamine; MurNAc, N-acetyl muramic acid; Rbo-P, ribitol phosphate; Rib-P, ribulose5-phosphate; UDP-Glc, uridine-5-diphosphate-glucose; UDP-GlcNAc, uridine-5-diphosphate-N-acetyl-glucosamine; UDP-ManNAc, uridine-5 diphosphate-N-acetylmannosamine.

ManNAc, respectively, from UDP-activated precursor molecules to C55-P (Soldo et al., 2002a; Ginsberg et al., 2006). UDP-ManNAc, which is otherwise not used in the S. aureus primary metabolism, is probably generated from UDP-GlcNAc by MnaA as in B. subtilis (Soldo et al., 2002b) and Listeria monocytogenes (Dubail et al., 2006). Incorporation of the preformed repeating units into WTA is mediated by a group of proteins, which comprise both, priming and polymerizing enzymes. The poly-Gro-P WTA of B. subtilis 168 depends on the primase TagB adding the rst Gro-P to the C55-Pcarried disaccharide linkage unit (Bhavsar et al., 2005; Ginsberg et al., 2006) and the polymerase TagF, which adds approximately 35 Gro-P units (Schertzer and Brown, 2003). The situation is more complicated in S. aureus Rbo-P WTA biosynthesis. Here the TagB reaction is followed by addition of only one additional Gro-P unit mediated by the TarF enzyme (Brown et al., 2008). The activated precursor molecule CDP-glycerol is generated by TagD (Park et al., 1993; Badurina et al., 2003). Subsequently, the TarL polymerase synthesizes the poly-Rbo-P (Brown et al., 2008; Meredith et al.,

2008; Pereira et al., 2008; Ishimoto and Strominger, 1966), while TarIJ are responsible for generating the precursor CDP-ribitol (Pereira and Brown, 2004). No Rbo-P primase seems to be involved in S. aureus, whereas such an enzyme (TarK) has been implicated in Rbo-P WTA biosynthesis in B. subtilis W23 (Meredith et al., 2008; Pereira et al., 2008). A major challenge in studying enzyme functions results from the fact that the three genes tarIJL involved in generation and incorporation of Rbo-P are duplicated in S. aureus (Qian et al., 2006). Recent studies indicate that the two functionally redundant TarL-like enzymes mediate the same types of reaction albeit leading to WTA of different chain length and electrophoretic mobility (Meredith et al., 2008; Pereira et al., 2008). Apparently, S. aureus can modulate its WTA structure according to bacterial density and environmental changes, since one of the tarL genes (also been named tarK) is repressed by the agr quorum-sensing system (Meredith et al., 2008). Eventually, the WTA polymers are translocated to the outer membrane leaet by an ABC transporter formed by TagG and TagH (Lazarevic and

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Karamata, 1995) and transferred from C55-P to the 6-OH group of MurNAc in the PG by a yet unknown enzyme (Mauck and Glaser, 1972; Fiedler et al., 1974; Bhavsar and Brown, 2006). The Gro-P repeating units of LTA are not derived from a nucleotide-activated precursor but from phosphatidylglycerol, a major constituent of bacterial membranes (Glaser and Lindsay, 1974) (Fig. 2B). LTA is polymerized directly on the glycolipid serving as the membrane anchor for LTA instead of C55-P, which is the second major difference between LTA and WTA biosynthesis (Koch et al., 1984; Fischer, 1988). The glycolipid is generated by the YpfP enzyme, which adds two glucose residues from UDPglucose to diacylglycerol (Jorasch et al., 1998, 2000; Kiriukhin et al., 2001). A membrane protein encoded by the ltaA gene is required for efcient LTA biosynthesis and is thought to be a ippase that translocates the glycolipid from the inner to the outer leaet of the cytoplasmic membrane (Grundling and Schneewind, 2007a). Recently, the LTA polymerase LtaS has been identied and cloned, which utilizes Gro-P units from phosphatidylglycerol to synthesize the LTA polymer at the outer surface of the cytoplasmic membrane (Grundling and Schneewind, 2007b). Amazingly, deletion of ypfP does not block biosynthesis of LTA but leads to synthesis of LTA polymer that is attached to diacylglycerol (Kiriukhin et al., 2001; Fedtke et al., 2007). For unknown reasons, ypfP mutants can produce either unaltered or strongly reduced amounts of LTA compared to the wild-type strains depending on the S. aureus strain background (Fedtke et al., 2007). A very constant trait of most TAs is the modication with D-alanine after biosynthesis of the polymers is completed. D-Alanine can be repeatedly incorporated into a given molecule, since the D-alanine esters are rather labile and get easily lost (Koch et al., 1985). The dltABCD genes responsible for D-alanine activation and incorporation into WTA and LTA are highly conserved and always seem to form an operon (Debabov et al., 1996; Neuhaus et al., 1996; Neuhaus and Baddiley, 2003). TA net charge is strongly affected by this modication (Peschel et al., 1999). The dlt operons of S. aureus and S. epidermidis are controlled by cations (Koprivnjak et al., 2006) and in response to CAMP challenge via the GraXRS (also named ApsXRS) regulatory system, which is in accord with the crucial role of TA alanylation in bacterial resistance to CAMPs (Li et al., 2007a, b; Bera et al., 2007; Kraus et al., 2008). Among the four dlt genes, the gene dltC encodes a D-alanyl carrier protein (Dcp) and dltA a ligase (Dcl), which catalyze the formation of D-alanyl-Dcp in the cytoplasm (Fig. 2C). DltB, an integral membrane protein and DltD, a membrane-tethered hydrophilic protein, seem to be required for the translocation and incorporation of D-alanine into TAs (Debabov et al., 2000; Neuhaus and Baddiley, 2003).

L. monocytogenes (Uchikawa et al., 1986), and Streptococcus pneumoniae (Fischer et al., 1993; Baur et al., 2009) in addition to S. aureus. Recently described crystal structures of proteins such as TagD (Fong et al., 2006), TarI (Baur et al., 2009), LtaS (Schirner et al., 2009; Lu et al., 2009), DltA (Du et al., 2008; Osman et al., 2009), and the Bacillus anthracis MnaA homolog (Velloso et al., 2008) represent the basis for rational drug design. Accordingly, specic inhibitors of DltA have recently been shown to render bacteria highly susceptible to CAMPs and cationic antibiotics (May et al., 2005) and to be very effective in clearing Gram-positive infections in vivo (Escaich et al., 2007). In addition, TAs have been shown to hold promise as targets for active or passive vaccines (Theilacker et al., 2004; Kumar et al., 2005; Weisman, 2007; Weisman et al., 2009). A broader view on the diversity and variability of TA structures can be achieved in the near future with improved glycobiochemical methods. The availability of genomic and metagenomic databases represents a valuable basis for predicting TA biosynthetic pathways by bioinformatic methods. Future studies will allow correlating structural features of TAs with certain functions in the physiology of cell envelope or bacteriahost interaction by making use of the increasing number of dened bacterial mutants with altered or lacking TAs. The controversial proinammatory capacity of LTA and the potential of zwitterionic TAs to activate specic T cells upon processing and presentation by MHC class II molecules are major open questions, which need to be approached. Furthermore, many host receptors recognizing and binding TAs remain to be identied. According to cell- and species-specic differences in expression of TA-binding molecules, it is tempting to assume that the enormous diversity of TA structures plays a role in bacterial cell and host tropism, in eluding the host immune system, and in adapting to varying environmental conditions.

Acknowledgments The authors acknowledge support by grants from the German Research Foundation (TR-SFB34, SFB766, FOR449, GRK685, SPP1130), the German Ministry of Education and Research (SkinStaph), and the IZKF program of the Medical Faculty, University of Tubingen. We apologize to those investigators whose work is not cited in this article. Unfortunately, space restrictions do not allow us to cite all of the relevant literature we refer to or all investigators who have contributed signicantly to this important and growing eld.

References Conclusions and perspectives The WTA biosynthetic enzymes seem to form a membraneassociated complex in B. subtilis, indicating that the biosynthesis of TAs is a highly organized process (Formstone et al., 2008). It can be assumed that the TA and PG biosynthetic machineries are coordinated in sophisticated ways, maybe in cooperation with cytoskeletal proteins and with the cell division apparatus (Schirner et al., 2009). How similar or different the rod-shaped B. subtilis and the spherical S. aureus are in this respect remains to be analyzed. While we are only beginning to understand the roles of WTA and LTA it is clear now that many of the biosynthetic enzymes represent attractive targets for new anti-infective agents. Recently, proteins involved in Rbo-P biosynthesis and incorporation have attracted particular interest, since WTA with Rbo-P residues are found in several human pathogens such as Staphylococcus saprophyticus (Schumacher-Perdreau et al., 1978),
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