Streptolysin O and its Co-Toxin NAD-glycohydrolase Protect Group A from Xenophagic Killing

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Group A Streptococcus (Streptococcus pyogenes or GAS) causes pharyngitis, severe invasive infections, and the post-infectious syndromes of glomerulonephritis and rheumatic fever. GAS can be internalized and killed by epithelial cells in vitro, a process that may contribute to local innate defense against pharyngeal infection. Secretion of the pore-forming toxin streptolysin O (SLO) by GAS has been reported to stimulate targeted autophagy (xenophagy) upon internalization of the bacteria by epithelial cells. Whereas this process was associated with killing of GAS in HeLa cells, studies in human keratinocytes found SLO production enhanced intracellular survival. To reconcile these conflicting observations, we now report in-depth investigation of xenophagy in response to GAS infection of human oropharyngeal keratinocytes, the predominant cell type of the pharyngeal epithelium. We found that SLO expression was associated with prolonged intracellular survival; unexpectedly, expression of the co-toxin NADase was required for this effect. Enhanced intracellular survival was lost upon deletion of NADase or inactivation of its enzymatic activity. Shortly after internalization of GAS by keratinocytes, SLO-mediated damage to the bacteria-containing vacuole resulted in exposure to the cytosol, ubiquitination of GAS and/or associated vacuolar membrane remnants, and engulfment of GAS in LC3-positive vacuoles. We also found that production of streptolysin S could mediate targeting of GAS to autophagosomes in the absence of SLO, a process accompanied by galectin 8 binding to damaged GAS-containing endosomes. Maturation of GAS-containing autophagosome-like vacuoles to degradative autolysosomes was prevented by SLO pore-formation and by SLO-mediated translocation of enzymatically active NADase into the keratinocyte cytosol. We conclude that SLO stimulates xenophagy in pharyngeal keratinocytes, but the coordinated action of SLO and NADase prevent maturation of GAS-containing autophagosomes, thereby prolonging GAS intracellular survival. This novel activity of NADase to block autophagic killing of GAS in pharyngeal cells may contribute to pharyngitis treatment failure, relapse, and chronic carriage.

Vyšlo v časopise: Streptolysin O and its Co-Toxin NAD-glycohydrolase Protect Group A from Xenophagic Killing. PLoS Pathog 9(6): e32767. doi:10.1371/journal.ppat.1003394
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1003394

Souhrn

Zdroje

1. Stevens DL (2000) Group A beta-hemolytic streptococci: virulence factors, pathogenesis, and spectrum of clinical infections. In: Stevens DL, Kaplan EL, editors. Streptococcal Infections: Clinical Aspects, Microbiology, and Molecular Pathogenesis. New York: Oxford University Press. pp. 19–36.

2. GerberMA, TanzRR, KabatW, BellGL, SiddiquiB, et al. (1999) Potential mechanisms for failure to eradicate group A streptococci from the pharynx. Pediatrics 104: 911–917.

3. PichicheroME, CaseyJR, MayesT, FrancisAB, MarsocciSM, et al. (2000) Penicillin failure in streptococcal tonsillopharyngitis: causes and remedies. Pediatr Infect Dis J 19: 917–923.

4. GrecoR, De MartinoL, DonnarummaG, ConteMP, SegantiL, et al. (1995) Invasion of cultured human cells by Streptococcus pyogenes. Res Microbiol 146: 551–560.

5. JadounJ, OzeriV, BursteinE, SkutelskyE, HanskiE, et al. (1998) Protein F1 is required for efficient entry of Streptococcus pyogenes into epithelial cells. J Infect Dis 178: 147–158.

6. LaPentaD, RubensC, ChiE, ClearyPP (1994) Group A streptococci efficiently invade human respiratory epithelial cells. Proc Natl Acad Sci USA 91: 12115–12119.

7. MolinariG, TalaySR, Valentin-WeigandP, RohdeM, ChhatwalGS (1997) The fibronectin-binding protein of Streptococcus pyogenes, SfbI, is involved in the internalization of group A streptococci by epithelial cells. Infect Immun 65: 1357–1363.

8. OsterlundA, EngstrandL (1995) Intracellular penetration and survival of Streptococcus pyogenes in respiratory epithelial cells in vitro. Acta Otolaryngologica 115: 685–688.

9. SchragerHM, RheinwaldJG, WesselsMR (1996) Hyaluronic acid capsule and the role of streptococcal entry into keratinocytes in invasive skin infection. J Clin Invest 98: 1954–1958.

10. OsterlundA, PopaR, NikkilaT, ScheyniusA, EngstrandL (1997) Intracellular reservoir of Streptococcus pyogenes in vivo: a possible explanation for recurrent pharyngotonsillitis. Laryngoscope 107: 640–647.

11. MarouniMJ, SelaS (2004) Fate of Streptococcus pyogenes and epithelial cells following internalization. J Med Microbiol 53: 1–7.

12. SpinaciC, MagiG, VaraldoPE, FacinelliB (2006) Persistence of erythromycin-resistant group a streptococci in cultured respiratory cells. Pediatr Infect Dis J 25: 880–883.

13. Cywes BentleyC, HakanssonA, ChristiansonJ, WesselsMR (2005) Extracellular group A Streptococcus induces keratinocyte apoptosis by dysregulating calcium signalling. Cell Microbiol 7: 945–955.

14. HakanssonA, BentleyCC, ShakhnovicEA, WesselsMR (2005) Cytolysin-dependent evasion of lysosomal killing. Proc Natl Acad Sci U S A 102: 5192–5197.

15. LogsdonLK, HakanssonAP, CortesG, WesselsMR (2011) Streptolysin O inhibits clathrin-dependent internalization of group A Streptococcus. mBio 2: e00332–00310.

16. NakagawaI, AmanoA, MizushimaN, YamamotoA, YamaguchiH, et al. (2004) Autophagy defends cells against invading group A Streptococcus. Science 306: 1037–1040.

17. SakuraiA, MaruyamaF, FunaoJ, NozawaT, AikawaC, et al. (2010) Specific behavior of intracellular Streptococcus pyogenes that has undergone autophagic degradation is associated with bacterial streptolysin O and host small G proteins Rab5 and Rab7. J Biol Chem 285: 22666–22675.

18. GilbertRJ (2010) Cholesterol-dependent cytolysins. Adv Exp Med Biol 677: 56–66.

19. HotzeEM, Wilson-KubalekE, FarrandAJ, BentsenL, ParkerMW, et al. (2012) Monomer-Monomer Interactions Propagate Structural Transitions Necessary for Pore Formation by the Cholesterol-dependent Cytolysins. J Biol Chem 287: 24534–24543.

20. MalleyR, HennekeP, MorseSC, CieslewiczMJ, LipsitchM, et al. (2003) Recognition of pneumolysin by Toll-like receptor 4 confers resistance to pneumococcal infection. Proc Natl Acad Sci U S A 100: 1966–1971.

21. ParkJM, NgVH, MaedaS, RestRF, KarinM (2004) Anthrolysin O and other gram-positive cytolysins are toll-like receptor 4 agonists. J Exp Med 200: 1647–1655.

22. PortnoyDA, AuerbuchV, GlomskiIJ (2002) The cell biology of Listeria monocytogenes infection: the intersection of bacterial pathogenesis and cell-mediated immunity. J Cell Biol 158: 409–414.

23. LevineB, MizushimaN, VirginHW (2011) Autophagy in immunity and inflammation. Nature 469: 323–335.

24. MizushimaN, KomatsuM (2011) Autophagy: renovation of cells and tissues. Cell 147: 728–741.

25. KimotoH, FujiiY, HiranoS, YokotaY, TaketoA (2006) Genetic and biochemical properties of streptococcal NAD-glycohydrolase inhibitor. J Biol Chem 281: 9181–9189.

26. MeehlMA, PinknerJS, AndersonPJ, HultgrenSJ, CaparonMG (2005) A Novel Endogenous Inhibitor of the Secreted Streptococcal NAD-Glycohydrolase. PLoS Pathog 1: e35.

27. BrickerAL, CywesC, AshbaughCD, WesselsMR (2002) NAD+-glycohydrolase acts as an intracellular toxin to enhance the extracellular survival of group A streptococci. Mol Microbiol 44: 257–269.

28. MaddenJC, RuizN, CaparonM (2001) Cytolysin-mediated translocation (CMT): a functional equivalent of type III secretion in gram-positive bacteria. Cell 104: 143–152.

29. MichosA, GryllosI, HakanssonA, SrivastavaA, KokkotouE, et al. (2006) Enhancement of streptolysin O activity and intrinsic cytotoxic effects of the group A streptococcal toxin, NAD-glycohydrolase. J Biol Chem 281: 8216–8223.

30. RiddleDJ, BessenDE, CaparonMG (2010) Variation in Streptococcus pyogenes NAD+ glycohydrolase is associated with tissue tropism. J Bacteriol 192: 3735–3746.

31. TatsunoI, SawaiJ, OkamotoA, MatsumotoM, MinamiM, et al. (2007) Characterization of the NAD-glycohydrolase in streptococcal strains. Microbiology 153: 4253–4260.

32. MagassaN, ChandrasekaranS, CaparonMG (2010) Streptococcus pyogenes cytolysin-mediated translocation does not require pore formation by streptolysin O. EMBO reports 11: 400–405.

33. AshbaughCD, WarrenHB, CareyVJ, WesselsMR (1998) Molecular analysis of the role of the group A streptococcal cysteine protease, hyaluronic acid capsule, and M protein in a murine model of human invasive soft-tissue infection. J Clin Invest 102: 550–560.

34. FujitaN, YoshimoriT (2011) Ubiquitination-mediated autophagy against invading bacteria. Current opinion in Cell Biology 23: 492–497.

35. RandowF (2011) How cells deploy ubiquitin and autophagy to defend their cytosol from bacterial invasion. Autophagy 7: 304–309.

36. Ginsberg I (1970) Streptolysin S. In: Montie TC, Kadis S, Ajil S, editors. Microbial toxins. New York: Academic Press. pp. 99–171.

37. ThurstonTL, WandelMP, von MuhlinenN, FoegleinA, RandowF (2012) Galectin 8 targets damaged vesicles for autophagy to defend cells against bacterial invasion. Nature 482: 414–418.

38. OliverMA, Garcia-ReyC, BoschR, AlbertiS (2007) Evaluation of the ability of erythromycin-resistant and -susceptible pharyngeal group A Streptococcus isolates from Spain to enter and persist in human keratinocytes. J Med Microbiol 56: 1485–1489.

39. BirminghamCL, CanadienV, KaniukNA, SteinbergBE, HigginsDE, et al. (2008) Listeriolysin O allows Listeria monocytogenes replication in macrophage vacuoles. Nature 451: 350–354.

40. BirminghamCL, SmithAC, BakowskiMA, YoshimoriT, BrumellJH (2006) Autophagy controls Salmonella infection in response to damage to the Salmonella-containing vacuole. J Biol Chem 281: 11374–11383.

41. RichKA, BurkettC, WebsterP (2003) Cytoplasmic bacteria can be targets for autophagy. Cell Microbiol 5: 455–468.

42. CywesC, StamenkovicI, WesselsMR (2000) CD44 as a receptor for colonization of the pharynx by group A Streptococcus. J Clin Invest 106: 995–1002.

43. RuizN, WangB, PentlandA, CaparonM (1998) Streptolysin O and adherence synergistically modulate proinflammatory responses of keratinocytes to group A streptococci. Mol Microbiol 27: 337–346.

44. ScottJR, GuenthnerPC, MaloneLM, FischettiVA (1986) Conversion of an M− group A streptococcus to M+ by transfer of a plasmid containing an M6 gene. J Exp Med 164: 1641–1651.

45. HillJE, WannamakerLW (1981) Identification of a lysin associated with a bacteriophage (A25) virulent for group A streptococci. J Bacteriol 145: 696–703.

46. Perez-CasalJ, PriceJA, MaguinE, ScottJR (1993) An M protein with a single C repeat prevents phagocytosis of Streptococcus pyogenes: use of a temperature-sensitive shuttle vector to deliver homologous sequences to the chromosome of S. pyogenes. Mol Microbiol 8: 809–819.

47. SierigG, CywesC, WesselsMR, AshbaughCD (2003) Cytotoxic effects of streptolysin O and streptolysin S enhance the virulence of poorly encapsulated group A streptococci. Infect Immun 71: 446–455.

48. LeBlancDJ, LeeLN, Abu-Al-JaibatA (1992) Molecular, genetic, and functional analysis of the basic replicon of pVA380-1, a plasmid of oral streptococcal origin. Plasmid 28: 130–145.

49. DicksonMA, HahnWC, InoY, RonfardV, WuJY, et al. (2000) Human keratinocytes that express hTERT and also bypass a p16INK4a-enforced mechanism that limits life span become immortal yet retain normal growth and differentiation characteristics. Mol Cell Biol 20: 1436–1447.

50. MorgensternJP, LandH (1990) Advanced mammalian gene transfer: high titre retroviral vectors with multiple drug selection markers and a complementary helper-free packaging cell line. Nucleic Acids Res 18: 3587–3596.