The Capsular Polysaccharide of Staphylococcus aureus Is Attached to Peptidoglycan by the LytR-CpsA-Psr (LCP) Family of Enzymes

Chan, Yvonne G. Y.; Kim, Hwan Keun; Schneewind, Olaf; Missiakas, Dominique

doi:10.1074/jbc.m114.567669

Cited by 88 publications

(94 citation statements)

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“…Scale bar ϭ 10 m. capsular polysaccharide. Nevertheless, each enzyme displays functional redundancy and catalyzes attachment of both WTA and polysaccharide to peptidoglycan (33 Mutations in B. anthracis lcpD or lcpB3 are associated with cell size and chain length phenotypes that have their underpinnings in SCWP assembly defects. In this regard, B. anthracis more closely resembles B. subtilis, in which mutations in individual lcp genes (tagT, tagU, and tagV) display discrete defects in teichoic acid attachment to peptidoglycan and in which the LCP enzymes TagT, TagU, and TagV assemble along MreB scaffolds (29,53), cytoskeletal elements that inform assembly and disassembly of peptidoglycan and thereby determine bacterial cell shape (54,55).…”

Section: Discussionmentioning

confidence: 99%

“…Of note, recent work implicated an LCP-like protein from A. orsi in the glycosylation of a sortase-anchored surface protein, although the chemical nature of the glycosyl modification is still unknown (35). S. aureus Newman variants lacking all three lcp genes (⌬lcp) cannot attach either WTA or the type 5 capsule, a polysaccharide with the repeat struc- (50), to peptidoglycan (32,33). Staphylococcal WTA synthesis requires tagO and tagA, whose products promote synthesis of murein linkage units (GlcNAc-ManNAc) on bactoprenol carriers (49,51,52).…”

Section: Discussionmentioning

confidence: 99%

“…Staphylococcal WTA synthesis requires tagO and tagA, whose products promote synthesis of murein linkage units (GlcNAc-ManNAc) on bactoprenol carriers (49,51,52). The tagO murein linkage pathway is dispensable for the synthesis of capsular polysaccharide, which also involves bactoprenol as the lipid carrier (33). When examined with single and double lcp mutants, S. aureus LcpA Sa , LcpB Sa , and LcpC Sa have been found to exert substrate preferences for WTA or Sterne (WT with empty vector) or lcpB3 or lcpD mutant strains containing empty vector or complementing plasmid were germinated in BHI broth, incubated for 8 h, fixed in formalin, and stained with polyclonal anti-BslR (A) or anti-BslU (B) rabbit antibodies.…”

Section: Discussionmentioning

confidence: 99%

“…The genome of Staphylococcus aureus contains three lcp genes, designated lcpA, lcpB, and lcpC (31,32). Staphylococcal mutants lacking all three lcp genes (⌬lcp), although viable, display growth defects, are unable to attach either WTA or capsular polysaccharide to pepti-doglycan, and release small amounts of these polymers into the extracellular medium (32)(33)(34). In Actinomyces oris, depletion of sortase A prevents bacterial growth and this phenotype can be suppressed by transposon insertions in a gene cluster encoding a LytR-CpsA-Psr (LCP)-like protein as well as with GspA (AcaA), a glycosylated surface protein substrate of sortase (35).…”

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LytR-CpsA-Psr Enzymes as Determinants of Bacillus anthracis Secondary Cell Wall Polysaccharide Assembly

et al. 2014

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bBacillus anthracis, the causative agent of anthrax, replicates as chains of vegetative cells by regulating the separation of septal peptidoglycan. Surface (S)-layer proteins and associated proteins (BSLs) function as chain length determinants and bind to the secondary cell wall polysaccharide (SCWP). In this study, we identified the B. anthracis lcpD mutant, which displays increased chain length and S-layer assembly defects due to diminished SCWP attachment to peptidoglycan. In contrast, the B. anthracis lcpB3 variant displayed reduced cell size and chain length, which could be attributed to increased deposition of BSLs. In other bacteria, LytR-CpsA-Psr (LCP) proteins attach wall teichoic acid (WTA) and polysaccharide capsule to peptidoglycan. B. anthracis does not synthesize these polymers, yet its genome encodes six LCP homologues, which, when expressed in S. aureus, promote WTA attachment. We propose a model whereby B. anthracis LCPs promote attachment of SCWP precursors to discrete locations in the peptidoglycan, enabling BSL assembly and regulated separation of septal peptidoglycan. Bacillus anthracis, the causative agent of anthrax, is a sporeforming, Gram-positive bacterium that germinates in host infected tissues and replicates as elongated chains of vegetative forms (1, 2). This unique growth pattern appears to be caused by the regulated separation of septal peptidoglycan, which generates bacterial chains whose mere size precludes clearance by host phagocytes (3-5). The genetic determinants for B. anthracis chain formation are conserved among pathogenic species of the Bacillus cereus group and, as demonstrated with B. cereus G9241, likely contribute to disease pathogenesis (6, 7). Hallmarks of pathogenic Bacillus species are virulence plasmids, pXO-1 and pXO-2 in B. anthracis (8, 9), providing for toxin production and capsulation (10, 11) as well as the chromosomally encoded surface (S)-layer locus (12). Two S-layer proteins of B. anthracis, Sap and EA1, are endowed with S-layer homology (SLH) domains, which retain these proteins in the bacterial envelope by binding to the secondary cell wall polysaccharide (SCWP) (13-15). S-layer protein crystallization domains, responsible for the spontaneous assembly of these polypeptides into a paracrystalline array (16), form a twodimensional lattice that can be thought of as bacterial integument (17)(18)(19).The structural genes of S-layer proteins, sap and eag, are flanked by genes encoding determinants for S-layer protein secretion (secA2 and slaP) (12) or pyruvylation (csaB) (14) as well as acetylation of the SCWP (patA1/2 and patB1/2) (20). CsaB-mediated pyruvylation of the terminal N-acetylmannosamine (ManNAc) of the SCWP (21), with the repeat struc- (22), is a prerequisite for the assembly of S-layer proteins and 22 B. anthracis S-layer-associated proteins (BSLs) (14). PatAB1/2-mediated acetylation of SCWP molecules affects the assembly of EA1 and of some but not all BSLs (20). Recent studies have begun to identify genes for SCWP synthesis, which, unlike py...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

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LytR-CpsA-Psr Enzymes as Determinants of Bacillus anthracis Secondary Cell Wall Polysaccharide Assembly

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“…We predict that specific LCP enzymes, catalysts of the phosphodiester linkage tethering teichoic acids and polysaccharides to peptidoglycan (20,51), also play a key role in the assembly of the SCWP at discrete sites. For example, tagOmediated SCWP attachment and subsequent assembly of Sap Slayers may require specific LCP enzymes from a cast of six different catalysts in B. anthracis (LcpB1-4, LcpC, and LcpD) (37), whereas tagO-independent attachment of the SCWP would involve others.…”

Section: Discussionmentioning

confidence: 99%

Bacillus anthracis tagO Is Required for Vegetative Growth and Secondary Cell Wall Polysaccharide Synthesis

et al. 2015

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Bacillus anthracis elaborates a linear secondary cell wall polysaccharide (SCWP) that retains surface (S)-layer and associated proteins via their S-layer homology (SLH) domains. The SCWP is comprised of trisaccharide repeats [¡4)-␤-ManNAc-(1¡4)-␤-GlcNAc-(1¡6)-␣-GlcNAc-(1¡]and tethered via acid-labile phosphodiester bonds to peptidoglycan. Earlier work identified UDP-GlcNAc 2-epimerases GneY (BAS5048) and GneZ (BAS5117), which act as catalysts of ManNAc synthesis, as well as a polysaccharide deacetylase (BAS5051), as factors contributing to SCWP synthesis. Here, we show that tagO (BAS5050), which encodes a UDP-N-acetylglucosamine:undecaprenyl-P N-acetylglucosaminyl 1-P transferase, the enzyme that initiates the synthesis of murein linkage units, is required for B. anthracis SCWP synthesis and S-layer assembly. Similar to gneY-gneZ mutants, B. anthracis strains lacking tagO cannot maintain cell shape or support vegetative growth. In contrast, mutations in BAS5051 do not affect B. anthracis cell shape, vegetative growth, SCWP synthesis, or S-layer assembly. These data suggest that TagO-mediated murein linkage unit assembly supports SCWP synthesis and attachment to the peptidoglycan via acid-labile phosphodiester bonds. Further, B. anthracis variants unable to synthesize SCWP trisaccharide repeats cannot sustain cell shape and vegetative growth. IMPORTANCEBacillus anthracis elaborates an SCWP to support vegetative growth and envelope assembly. Here, we show that some, but not all, SCWP synthesis is dependent on tagO-derived murein linkage units and subsequent attachment of SCWP to peptidoglycan. The data implicate secondary polymer modifications of peptidoglycan and subcellular distributions as a key feature of the cell cycle in Gram-positive bacteria and establish foundations for work on the molecular functions of the SCWP and on inhibitors with antibiotic attributes.T he cell wall envelope of Bacillus anthracis, the causative agent of anthrax, is comprised of peptidoglycan and its attached secondary cell wall polysaccharide (SCWP) (1). The SCWP retains two surface (S)-layer proteins, surface array protein (Sap) and extractable antigen 1 (EA1) (2), as well as 22 S-layer-associated proteins whose S-layer homology (SLH) domains associate with ketal-pyruvylated SCWP (3, 4). Unlike nonpathogenic Bacillus spp., for example, Bacillus subtilis and Bacillus cereus AHU1030, B. anthracis does not synthesize wall teichoic acid (WTA) (5, 6). Nevertheless, B. anthracis expresses functional tagO and tagA genes, whose products provide for the synthesis of murein linkage units [P-GlcNAc-(4¡1)-␤-ManNAc-R] that are phosphodiester linked to the C 6 -hydroxyl of N-acetylmuramic acid in the repeating disaccharide of peptidoglycan [MurNAc(P-GlcNAc-ManNAc-R)-GlcNAc] (4). In B. subtilis, -R represents wall teichoic acid, i.e., either polyglycerol-phosphate or polyribitol-phosphate (7). Here, we test the hypothesis that in B. anthracis the SCWP is attached via the murine linkage unit and that the tagO gene is required for vegetati...

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Membrane Translocation and Assembly of Sugar Polymer Precursors

Taylor

Huszczynski

Lam

2015

Current Topics in Microbiology and Immunology

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Bacterial polysaccharides play an essential role in cell viability, virulence, and evasion of host defenses. Although the polysaccharides themselves are highly diverse, the pathways by which bacteria synthesize these essential polymers are conserved in both Gram-negative and Gram-positive organisms. By utilizing a lipid linker, a series of glycosyltransferases and integral membrane proteins act in concert to synthesize capsular polysaccharide, teichoic acid, and teichuronic acid. The pathways used to produce these molecules are the Wzx/Wzy-dependent, the ABC-transporter-dependent, and the synthase-dependent pathways. This chapter will cover the initiation, synthesis of the various polysaccharides on the cytoplasmic face of the membrane using nucleotide sugar precursors, and export of the nascent chain from the cytoplasm to the extracellular milieu. As microbial glycobiology is an emerging field in Gram-positive bacteria research, parallels will be drawn to the more widely studied polysaccharide biosynthesis systems in Gram-negative species in order to provide greater understanding of these biologically significant molecules.

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The Capsular Polysaccharide of Staphylococcus aureus Is Attached to Peptidoglycan by the LytR-CpsA-Psr (LCP) Family of Enzymes

Cited by 88 publications

References 72 publications

LytR-CpsA-Psr Enzymes as Determinants of Bacillus anthracis Secondary Cell Wall Polysaccharide Assembly

LytR-CpsA-Psr Enzymes as Determinants of Bacillus anthracis Secondary Cell Wall Polysaccharide Assembly

Bacillus anthracis tagO Is Required for Vegetative Growth and Secondary Cell Wall Polysaccharide Synthesis

Membrane Translocation and Assembly of Sugar Polymer Precursors

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