Abstract
Knockout of lprG results in decreased virulence of Mycobacterium tuberculosis (MTB) in mice. MTB lipoprotein LprG has TLR2 agonist activity, which is thought to be dependent on its N-terminal triacylation. Unexpectedly, here we find that nonacylated LprG retains TLR2 activity. Moreover, we show LprG association with triacylated glycolipid TLR2 agonists lipoarabinomannan, lipomannan and phosphatidylinositol mannosides (which share core structures). Binding of triacylated species was specific to LprG (not LprA) and increased LprG TLR2 agonist activity; conversely, association of glycolipids with LprG enhanced their recognition by TLR2. The crystal structure of LprG in complex with phosphatidylinositol mannoside revealed a hydrophobic pocket that accommodates the three alkyl chains of the ligand. In conclusion, we demonstrate a glycolipid binding function of LprG that enhances recognition of triacylated MTB glycolipids by TLR2 and may affect glycolipid assembly or transport for bacterial cell wall biogenesis.
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References
Flynn, J.L. & Chan, J. Immunology of tuberculosis. Annu. Rev. Immunol. 19, 93–129 (2001).
Janeway, C.A. Jr. Approaching the asymptote? Evolution and revolution in immunology. Cold Spring Harb. Symp. Quant. Biol. 54, 1–13 (1989).
Chan, J. et al. Microbial glycolipids: possible virulence factors that scavenge oxygen radicals. Proc. Natl. Acad. Sci. USA 86, 2453–2457 (1989).
Fratti, R.A., Chua, J., Vergne, I. & Deretic, V. Mycobacterium tuberculosis glycosylated phosphatidylinositol causes phagosome maturation arrest. Proc. Natl. Acad. Sci. USA 100, 5437–5442 (2003).
Vergne, I. et al. Mycobacterium tuberculosis phagosome maturation arrest: mycobacterial phosphatidylinositol analog phosphatidylinositol mannoside stimulates early endosomal fusion. Mol. Biol. Cell 15, 751–760 (2004).
Cole, S.T. et al. Deciphering the biology of Mycobacterium tuberculosis from the complete genome sequence. Nature 393, 537–544 (1998).
Sutcliffe, I.C. & Harrington, D.J. Lipoproteins of Mycobacterium tuberculosis: an abundant and functionally diverse class of cell envelope components. FEMS Microbiol. Rev. 28, 645–659 (2004).
Bigi, F. et al. The knockout of the lprG-Rv1410 operon produces strong attenuation of Mycobacterium tuberculosis . Microbes Infect. 6, 182–187 (2004).
Rengarajan, J., Bloom, B.R. & Rubin, E.J. Genome-wide requirements for Mycobacterium tuberculosis adaptation and survival in macrophages. Proc. Natl. Acad. Sci. USA 102, 8327–8332 (2005).
Sassetti, C.M. & Rubin, E.J. Genetic requirements for mycobacterial survival during infection. Proc. Natl. Acad. Sci. USA 100, 12989–12994 (2003).
Farrow, M.F. & Rubin, E.J. Function of a mycobacterial major facilitator superfamily pump requires a membrane-associated lipoprotein. J. Bacteriol. 190, 1783–1791 (2008).
Sulzenbacher, G. et al. LppX is a lipoprotein required for the translocation of phthiocerol dimycocerosates to the surface of Mycobacterium tuberculosis . EMBO J. 25, 1436–1444 (2006).
Jones, B.W. et al. Different Toll-like receptor agonists induce distinct macrophage responses. J. Leukoc. Biol. 69, 1036–1044 (2001).
Gilleron, M., Quesniaux, V.F. & Puzo, G. Acylation state of the phosphatidyl inositol hexamannosides from Mycobacterium bovis BCG and Mycobacterium tuberculosis H37Rv and its implication in TLR response. J. Biol. Chem. 278, 29880–29889 (2003).
Elass, E. et al. Mycobacterial lipomannan induces matrix metalloproteinase-9 expression in human macrophagic cells through a toll-like receptor 1 (TLR1)/TLR2- and CD14-dependent mechanism. Infect. Immun. 73, 7064–7068 (2005).
Tapping, R.I. & Tobias, P.S. Mycobacterial lipoarabinomannan mediates physical interactions between TLR1 and TLR2 to induce signaling. J. Endotoxin Res. 9, 264–268 (2003).
Bhatt, K. & Salgame, P. Host innate immune response to Mycobacterium tuberculosis . J. Clin. Immunol. 27, 347–362 (2007).
Pai, R.K., Convery, M., Hamilton, T.A., Boom, W.H. & Harding, C.V. Inhibition of IFN-γ-induced class II transactivator expression by a 19-kDa lipoprotein from Mycobacterium tuberculosis: a potential mechanism for immune evasion. J. Immunol. 171, 175–184 (2003).
Noss, E.H. et al. Toll-like receptor 2-dependent inhibition of macrophage class II MHC expression and antigen processing by 19 kD lipoprotein of Mycobacterium tuberculosis . J. Immunol. 167, 910–918 (2001).
Brightbill, H.D. et al. Host defense mechanisms triggered by microbial lipoproteins through toll-like receptors. Science 285, 732–736 (1999).
Takeuchi, O. et al. Cutting edge: role of Toll-like receptor 1 in mediating immune response to microbial lipoproteins. J. Immunol. 169, 10–14 (2002).
Nigou, J. et al. Mannan chain length controls lipoglycans signaling via and binding to TLR2. J. Immunol. 180, 6696–6702 (2008).
Pecora, N.D., Gehring, A.J., Canaday, D.H., Boom, W.H. & Harding, C.V. Mycobacterium tuberculosis LprA is a lipoprotein agonist of TLR2 that regulates innate immunity and APC function. J. Immunol. 177, 422–429 (2006).
Gehring, A.J., Dobos, K.M., Belisle, J.T., Harding, C.V. & Boom, W.H. Mycobacterium tuberculosis LprG (Rv1411c): A novel TLR-2 ligand that inhibits human macrophage class II MHC antigen processing. J. Immunol. 173, 2660–2668 (2004).
Jung, S.B. et al. The mycobacterial 38-kilodalton glycolipoprotein antigen activates the mitogen-activated protein kinase pathway and release of proinflammatory cytokines through Toll-like receptors 2 and 4 in human monocytes. Infect. Immun. 74, 2686–2696 (2006).
Jin, M.S. et al. Crystal structure of the TLR1–TLR2 heterodimer induced by binding of a tri-acylated lipopeptide. Cell 130, 1071–1082 (2007).
Rezwan, M., Grau, T., Tschumi, A. & Sander, P. Lipoprotein synthesis in mycobacteria. Microbiology 153, 652–658 (2007).
Nigou, J., Gilleron, M. & Puzo, G. Lipoarabinomannans: characterization of the multiacylated forms of the phosphatidyl-myo-inositol anchor by NMR spectroscopy. Biochem. J. 337, 453–460 (1999).
Bigi, F. et al. The gene encoding P27 lipoprotein and a putative antibiotic-resistance gene form an operon in Mycobacterium tuberculosis and Mycobacterium bovis . Microbiology 146, 1011–1018 (2000).
Gilleron, M., Nigou, J., Nicolle, D., Quesniaux, V. & Puzo, G. The acylation state of mycobacterial lipomannans modulates innate immunity response through toll-like receptor 2. Chem. Biol. 13, 39–47 (2006).
Kang, J.Y. et al. Recognition of lipopeptide patterns by Toll-like receptor 2-Toll-like receptor 6 heterodimer. Immunity 31, 873–884 (2009).
Berg, S., Kaur, D., Jackson, M. & Brennan, P.J. The glycosyltransferases of Mycobacterium tuberculosis—roles in the synthesis of arabinogalactan, lipoarabinomannan, and other glycoconjugates. Glycobiology 17, 35R–56R (2007).
Finberg, R.W., Re, F., Popova, L., Golenbock, D.T. & Kurt-Jones, E.A. Cell activation by Toll-like receptors: role of LBP and CD14. J. Endotoxin Res. 10, 413–418 (2004).
Jiang, Z. et al. CD14 is required for MyD88-independent LPS signaling. Nat. Immunol. 6, 565–570 (2005).
Sklar, M.D., Tereba, A., Chen, B.D. & Walker, W.S. Transformation of mouse bone marrow cells by transfection with a human oncogene related to c-myc is associated with the endogenous production of macrophage colony stimulating factor 1. J. Cell. Physiol. 125, 403–412 (1985).
Flo, T.H. et al. Involvement of toll-like receptor (TLR) 2 and TLR4 in cell activation by mannuronic acid polymers. J. Biol. Chem. 277, 35489–35495 (2002).
Latz, E. et al. Lipopolysaccharide rapidly traffics to and from the Golgi apparatus with the toll-like receptor 4-MD-2–CD14 complex in a process that is distinct from the initiation of signal transduction. J. Biol. Chem. 277, 47834–47843 (2002).
Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).
Vonrhein, C., Blanc, E., Roversi, P. & Bricogne, G. Automated structure solution with autoSHARP. Methods Mol. Biol. 364, 215–230 (2007).
Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).
Adams, P.D. et al. PHENIX: building new software for automated crystallographic structure determination. Acta Crystallogr. D Biol. Crystallogr. 58, 1948–1954 (2002).
Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).
Pettersen, E.F. et al. UCSF Chimera–a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).
Dundas, J. et al. CASTp: computed atlas of surface topography of proteins with structural and topographical mapping of functionally annotated residues. Nucleic Acids Res. 34, W116–8 (2006).
Acknowledgements
We thank V. Anderson and L. Sweet for advice and assistance with MS, A.G. Hise (Case Western Reserve Univ.) and A. Shizuo (Osaka Univ.) for bone marrow from TLR1−/− TLR6−/− mice, X. Ding, N. Nagy and K. Daniel for technical assistance, T. Musa for comments on the manuscript and K. Dobos-Elder and J. Belisle (Colorado State Univ.) for MTB lysates, antibodies and the pVV16 vector. This work was supported by US National Institutes of Health (NIH) grants AI035726, AI034343 and AI069085 to C.V.H., HL055967 and AI027243 to W.H.B., AI071155 and AI049313 to D.B.M. and AI068135 to J.C.S., the Robert A. Welch Foundation (J.C.S.), the Irvington Institute Fellowship Program of the Cancer Research Institute (C.S.), American Lung Association grant RG48786N (R.E.R.) and the Burroughs Wellcome Fund for Translational Research (D.B.M. and C.S.). Core facilities of the Case Western Reserve University Center for AIDS Research were supported by NIH grant AI067093.
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M.G.D., H.-C.T., N.D.P., T.-Y.C., A.R.A., S.S., R.E.R. and C.S. designed, performed and interpreted experiments and prepared the manuscript; D.B.M., W.H.B., J.C.S. and C.V.H. designed and interpreted experiments and prepared the manuscript.
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Drage, M., Tsai, HC., Pecora, N. et al. Mycobacterium tuberculosis lipoprotein LprG (Rv1411c) binds triacylated glycolipid agonists of Toll-like receptor 2. Nat Struct Mol Biol 17, 1088–1095 (2010). https://doi.org/10.1038/nsmb.1869
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DOI: https://doi.org/10.1038/nsmb.1869
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