The pH-Responsive PacC Transcription Factor of Governs Epithelial Entry and Tissue Invasion during Pulmonary Aspergillosis

English version České info

Inhaled spores of the pathogenic mould Aspergillus fumigatus cause fungal lung infections in humans having immune defects. A. fumigatus spores germinate within the immunocompromised lung, producing invasively growing, elongated cells called hyphae. Hyphae degrade the surrounding pulmonary tissue, a process thought to be caused by secreted fungal enzymes; however, A. fumigatus mutants lacking one or more protease activities retain fully invasive phenotypes in mouse models of disease. Here we report the first discovery of a non-invasive A. fumigatus mutant, which lacks a pH-responsive transcription factor PacC. Using global transcriptional profiling of wild type and mutant isolates, and in vitro pulmonary invasion assays, we established that loss of PacC leads to a compound non-invasive phenotype characterised by deficits in both contact-mediated epithelial entry and protease expression. Consistent with an important role for epithelial entry in promoting invasive disease in mammalian tissues, PacC mutants remain surface-localised on mammalian epithelia, both in vitro and in vivo. Our study sets a new precedent for involvement of both host and pathogen activities in promoting epithelial invasion by A. fumigatus and supports a model wherein fungal protease activity acting subsequently to, or in parallel with, host-mediated epithelial entry provides the mechanistic basis for tissue invasion.

Vyšlo v časopise: The pH-Responsive PacC Transcription Factor of Governs Epithelial Entry and Tissue Invasion during Pulmonary Aspergillosis. PLoS Pathog 10(10): e32767. doi:10.1371/journal.ppat.1004413
Kategorie: Research Article
prolekare.web.journal.doi_sk: https://doi.org/10.1371/journal.ppat.1004413

Souhrn

Zdroje

1. BrownGD, DenningDW, GowNA, LevitzSM, NeteaMG, et al. (2012) Hidden killers: human fungal infections. Sci Transl Med 4 165: 165rv13.

2. BaddleyJW, AndesDR, MarrKA, KontoyiannisDP, AlexanderBD, et al. (2010) Factors associated with mortality in transplant patients with invasive aspergillosis. Clin Infect Dis 50 12: 1559–67.

3. KontoyiannisDP, MarrKA, ParkBJ, AlexanderBD, AnaissieEJ, et al. (2010) Prospective surveillance for invasive fungal infections in hematopoietic stem cell transplant recipients, 2001-2006: overview of the Transplant-Associated Infection Surveillance Network (TRANSNET) Database. Clin Infect Dis 50 8: 1091–100.

4. PappasPG, AlexanderBD, AndesDR, HadleyS, KauffmanCA, et al. (2010) Invasive fungal infections among organ transplant recipients: results of the Transplant-Associated Infection Surveillance Network (TRANSNET). Clin Infect Dis 50 8: 1101–11.

5. DenningDW, PleuvryA, ColeDC (2011) Global burden of chronic pulmonary aspergillosis as a sequel to pulmonary tuberculosis. Bull World Health Organ 89 12: 864–72.

6. MorganJ, WannemuehlerKA, MarrKA, HadleyS, KontoyiannisDP, et al. (2005) Incidence of invasive aspergillosis following hematopoietic stem cell and solid organ transplantation: interim results of a prospective multicenter surveillance program. Med Mycol 43 Suppl 1 S49–58.

7. AmitaniR, TaylorG, ElezisEN, Llewellyn-JonesC, MitchellJ, et al. (1995) Purification and characterization of factors produced by Aspergillus fumigatus which affect human ciliated respiratory epithelium. Infect Immun 63 9: 3266–71.

8. KoganTV, JadounJ, MittelmanL, HirschbergK, OsherovN (2004) Involvement of secreted Aspergillus fumigatus proteases in disruption of the actin fiber cytoskeleton and loss of focal adhesion sites in infected A549 lung pneumocytes. J Infect Dis 189 11: 1965–73.

9. SharonH, HagagS, OsherovN (2009) Transcription factor PrtT controls expression of multiple secreted proteases in the human pathogenic mold Aspergillus fumigatus. Infect Immun 77 9: 4051–60.

10. BergmannA, HartmannT, CairnsT, BignellEM, KrappmannS (2009) A regulator of Aspergillus fumigatus extracellular proteolytic activity is dispensable for virulence. Infect Immun 77 9: 4041–50.

11. HartmannT, CairnsTC, OlbermannP, MorschhauserJ, BignellEM, et al. (2011) Oligopeptide transport and regulation of extracellular proteolysis are required for growth of Aspergillus fumigatus on complex substrates but not for virulence. Mol Microbiol 82 4: 917–35.

12. Ibrahim-GranetO, D'EnfertC (1997) The Aspergillus fumigatus mepB gene encodes an 82 kDa intracellular metalloproteinase structurally related to mammalian thimet oligopeptidases. Microbiology 143 (Pt 7) 2247–53.

13. Jaton-OgayK, ParisS, HuerreM, QuadroniM, FalchettoR, et al. (1994) Cloning and disruption of the gene encoding an extracellular metalloprotease of Aspergillus fumigatus. Mol Microbiol 14 5: 917–28.

14. ReichardU, MonodM, OddsF, RuchelR (1997) Virulence of an aspergillopepsin-deficient mutant of Aspergillus fumigatus and evidence for another aspartic proteinase linked to the fungal cell wall. J Med Vet Mycol 35 3: 189–96.

15. TangCM, CohenJ, KrauszT, Van NoordenS, HoldenDW (1993) The alkaline protease of Aspergillus fumigatus is not a virulence determinant in two murine models of invasive pulmonary aspergillosis. Infect Immun 61 5: 1650–6.

16. DeHartDJ, AgwuDE, JulianNC, WashburnRG (1997) Binding and germination of Aspergillus fumigatus conidia on cultured A549 pneumocytes. J Infect Dis 175 1: 146–50.

17. ParisS, Boisvieux-UlrichE, CrestaniB, HoucineO, TaramelliD, et al. (1997) Internalization of Aspergillus fumigatus conidia by epithelial and endothelial cells. Infect Immun 65 4: 1510–4.

18. WasylnkaJA, MooreMM (2002) Uptake of Aspergillus fumigatus conidia by phagocytic and nonphagocytic cells in vitro: quantitation using strains expressing green fluorescent protein. Infect Immun 70 6: 3156–63.

19. GomezP, HackettTL, MooreMM, KnightDA, TebbuttSJ (2010) Functional genomics of human bronchial epithelial cells directly interacting with conidia of Aspergillus fumigatus. BMC Genomics 11 358.

20. WasylnkaJA, MooreMM (2003) Aspergillus fumigatus conidia survive and germinate in acidic organelles of A549 epithelial cells. J Cell Sci 116 Pt 8: 1579–87.

21. OsherovN (2012) Interaction of the pathogenic mold Aspergillus fumigatus with lung epithelial cells. Front Microbiol 3 346.

22. ChaudharyN, DattaK, AskinFB, StaabJF, MarrKA (2012) Cystic fibrosis transmembrane conductance regulator regulates epithelial cell response to Aspergillus and resultant pulmonary inflammation. Am J Respir Crit Care Med 185 3: 301–10.

23. CaddickMX, BrownleeAG, ArstHNJr (1986) Regulation of gene expression by pH of the growth medium in Aspergillus nidulans. Mol Gen Genet 203 2: 346–53.

24. DavisD, WilsonRB, MitchellAP (2000) RIM101-dependent and-independent pathways govern pH responses in Candida albicans. Mol Cell Biol 20 3: 971–8.

25. DavisD, EdwardsJEJr, MitchellAP, IbrahimAS (2000) Candida albicans RIM101 pH response pathway is required for host-pathogen interactions. Infect Immun 68 10: 5953–9.

26. LiW, MitchellA (1997) Proteolytic activation of Rim1p, a positive regulator of yeast sporulation and invasive growth. Genetics 145 1: 63.

27. TilburnJ, SarkarS, WiddickDA, EspesoEA, OrejasM, et al. (1995) The Aspergillus PacC zinc finger transcription factor mediates regulation of both acid- and alkaline-expressed genes by ambient pH. EMBO J 14 4: 779–90.

28. DavisD (2003) Adaptation to environmental pH in Candida albicans and its relation to pathogenesis. Current genetics 44 1: 1–7.

29. BignellE, Negrete-UrtasunS, CalcagnoA, HaynesK, ArstHJr, et al. (2005) The Aspergillus pH-responsive transcription factor PacC regulates virulence. Molecular microbiology 55 4: 1072–1084.

30. McDonaghA, FedorovaND, CrabtreeJ, YuY, KimS, et al. (2008) Sub-Telomere Directed Gene Expression during Initiation of Invasive Aspergillosis. PLoS Pathog 4 9: e1000154.

31. PeñalvaM, TilburnJ, BignellE, ArstHJr (2008) Ambient pH gene regulation in fungi: making connections. Trends in microbiology 16 6: 291–300.

32. GrahlN, PuttikamonkulS, MacdonaldJM, GamcsikMP, NgoLY, et al. (2011) In vivo hypoxia and a fungal alcohol dehydrogenase influence the pathogenesis of invasive pulmonary aspergillosis. PLoS Pathog 7 7: e1002145.

33. AmichJ, VicentefranqueiraR, MelladoE, Ruiz-CarmuegaA, LealF, et al. (2014) The ZrfC alkaline zinc transporter is required for Aspergillus fumigatus virulence and its growth in the presence of the Zn/Mn-chelating protein calprotectin. Cell Microbiol 16 4: 548–64.

34. LewisRE, WiederholdNP (2005) Murine model of invasive aspergillosis. Methods Mol Med 118 129–42.

35. BezerraLML, FillerSG (2004) Interactions of Aspergillus fumigatus with endothelial cells: internalization, injury, and stimulation of tissue factor activity. Blood 103 6: 2143–2149.

36. MelladoE, Aufauvre-BrownA, GowNA, HoldenDW (1996) The Aspergillus fumigatus chsC and chsG genes encode class III chitin synthases with different functions. Mol Microbiol 20 3: 667–79.

37. ValianteV, HeinekampT, JainR, HartlA, BrakhageAA (2008) The mitogen-activated protein kinase MpkA of Aspergillus fumigatus regulates cell wall signaling and oxidative stress response. Fungal Genet Biol 45 5: 618–27.

38. DichtlK, HelmschrottC, DirrF, WagenerJ (2012) Deciphering cell wall integrity signalling in Aspergillus fumigatus: identification and functional characterization of cell wall stress sensors and relevant Rho GTPases. Mol Microbiol 83 3: 506–19.

39. CheungAL, YingP, FischettiVA (1991) A method to detect proteinase activity using unprocessed X-ray films. Anal Biochem 193 1: 20–3.

40. BrownGD, TaylorPR, ReidDM, WillmentJA, WilliamsDL, et al. (2002) Dectin-1 is a major β-glucan receptor on macrophages. J Exp Med 196 3: 407–12.

41. BrownGD, HerreJ, WilliamsDL, WillmentJA, MarshallAS, et al. (2003) Dectin-1 mediates the biological effects of β-glucans. J Exp Med 197 9: 1119–24.

42. BrownGD, GordonS (2001) Immune recognition. A new receptor for beta-glucans. Nature 413 6851: 36–7.

43. RandTG, SunM, GilyanA, DowneyJ, MillerJD (2010) Dectin-1 and inflammation-associated gene transcription and expression in mouse lungs by a toxic (1,3)-β-D glucan. Arch Toxicol 84 3: 205–20.

44. HanX, YuR, ZhenD, TaoS, SchmidtM, et al. (2011) beta-1,3-Glucan-induced host phospholipase D activation is involved in Aspergillus fumigatus internalization into type II human pneumocyte A549 cells. PLoS One 6 7: e21468.

45. GrahamLM, TsoniSV, WillmentJA, WilliamsDL, TaylorPR, et al. (2006) Soluble Dectin-1 as a tool to detect β-glucans. J Immunol Methods 314 1–2: 164–9.

46. Sousa MdaG, ReidDM, SchweighofferE, TybulewiczV, RulandJ, et al. (2011) Restoration of pattern recognition receptor costimulation to treat chromoblastomycosis, a chronic fungal infection of the skin. Cell Host Microbe 9 5: 436–43.

47. RappleyeCA, EissenbergLG, GoldmanWE (2007) Histoplasma capsulatum α-(1,3)-glucan blocks innate immune recognition by the β-glucan receptor. Proc Natl Acad Sci U S A 104 4: 1366–70.

48. SorciG, GiovanniniG, RiuzziF, BonifaziP, ZelanteT, et al. (2011) The danger signal S100B integrates pathogen- and danger-sensing pathways to restrain inflammation. PLoS Pathog 7 3: e1001315.

49. NobileCJ, SolisN, MyersCL, FayAJ, DeneaultJS, et al. (2008) Candida albicans transcription factor Rim101 mediates pathogenic interactions through cell wall functions. Cell Microbiol 10 11: 2180–96.

50. O'MearaTR, NortonD, PriceMS, HayC, ClementsMF, et al. (2010) Interaction of Cryptococcus neoformans Rim101 and protein kinase A regulates capsule. PLoS Pathog 6 2: e1000776.

51. O'MearaTR, HolmerSM, SelvigK, DietrichF, AlspaughJA (2013) Cryptococcus neoformans Rim101 is associated with cell wall remodeling and evasion of the host immune responses. MBio 4 1.

52. O'MearaTR, XuW, SelvigKM, O'MearaMJ, MitchellAP, et al. (2014) The Cryptococcus neoformans Rim101 Transcription Factor Directly Regulates Genes Required for Adaptation to the Host. Molecular and Cellular Biology 34 4: 673–684.

53. EUCAST (2008) Subcommittee on Antifungal Susceptibility Testing (AFST) of the ESCMID European Committee for Antimicrobial Susceptibility Testing (EUCAST), EUCAST Technical Note on the method for the determination of broth dilution minimum inhibitory concentrations of antifungal agents for conidia-forming moulds. Clin Microbiol Infect 14 10: 982–4.

54. EisendleM, ObereggerH, ButtingerR, IllmerP, HaasH (2004) Biosynthesis and uptake of siderophores is controlled by the PacC-mediated ambient-pH regulatory system in Aspergillus nidulans. Eukaryotic cell 3 2: 561.

55. NiermanWC, PainA, AndersonMJ, WortmanJR, KimHS, et al. (2005) Genomic sequence of the pathogenic and allergenic filamentous fungus Aspergillus fumigatus. Nature 438 7071: 1151–6.

56. LatgeJP (1999) Aspergillus fumigatus and aspergillosis. Clin Microbiol Rev 12 2: 310–50.

57. DagenaisTR, KellerNP (2009) Pathogenesis of Aspergillus fumigatus in Invasive Aspergillosis. Clin Microbiol Rev 22 3: 447–65.

58. KolattukudyPE, LeeJD, RogersLM, ZimmermanP, CeselskiS, et al. (1993) Evidence for possible involvement of an elastolytic serine protease in aspergillosis. Infect Immun 61 6: 2357–68.

59. MarkaryanA, MorozovaI, YuH, KolattukudyPE (1994) Purification and characterization of an elastinolytic metalloprotease from Aspergillus fumigatus and immunoelectron microscopic evidence of secretion of this enzyme by the fungus invading the murine lung. Infect Immun 62 6: 2149–57.

60. SharonH, AmarD, LevdanskyE, MircusG, ShadkchanY, et al. (2011) PrtT-regulated proteins secreted by Aspergillus fumigatus activate MAPK signaling in exposed A549 lung cells leading to necrotic cell death. PLoS One 6 3: e17509.

61. MonodM, ParisS, SarfatiJ, Jaton-OgayK, AveP, et al. (1993) Virulence of alkaline protease-deficient mutants of Aspergillus fumigatus. FEMS Microbiol Lett 106 1: 39–46.

62. ParkH, MyersCL, SheppardDC, PhanQT, SanchezAA, et al. (2005) Role of the fungal Ras-protein kinase A pathway in governing epithelial cell interactions during oropharyngeal candidiasis. Cell Microbiol 7 4: 499–510.

63. PhanQT, MyersCL, FuY, SheppardDC, YeamanMR, et al. (2007) Als3 is a Candida albicans invasin that binds to cadherins and induces endocytosis by host cells. PLoS Biol 5 3: e64.

64. SunJN, SolisNV, PhanQT, BajwaJS, KashlevaH, et al. (2010) Host cell invasion and virulence mediated by Candida albicans Ssa1. PLoS Pathog 6 11: e1001181.

65. WächtlerB, CitiuloF, JablonowskiN, FörsterS, DalleF, et al. (2012) Candida albicans-Epithelial Interactions: Dissecting the Roles of Active Penetration, Induced Endocytosis and Host Factors on the Infection Process. PLoS One 7 5: e36952.

66. MelkamuT, SquillaceD, KitaH, O'GradySM (2009) Regulation of TLR2 expression and function in human airway epithelial cells. J Membr Biol 229 2: 101–13.

67. LeeHM, YukJM, ShinDM, JoEK (2009) Dectin-1 is inducible and plays an essential role for mycobacteria-induced innate immune responses in airway epithelial cells. J Clin Immunol 29 6: 795–805.

68. LeeHM, ShinDM, ChoiDK, LeeZW, KimKH, et al. (2009) Innate immune responses to Mycobacterium ulcerans via toll-like receptors and dectin-1 in human keratinocytes. Cell Microbiol 11 4: 678–92.

69. VillarCC, KashlevaH, NobileCJ, MitchellAP, Dongari-BagtzoglouA (2007) Mucosal tissue invasion by Candida albicans is associated with E-cadherin degradation, mediated by transcription factor Rim101p and protease Sap5p. Infect Immun 75 5: 2126–35.

70. PontecorvoG, RoperJA, HemmonsLM, MacdonaldKD, BuftonAW (1953) The genetics of Aspergillus nidulans. Adv Genet 5 141–238.

71. DeightonFJ, HallNK, LarshHW (1979) Merthiolate treatment of pathogenic fungi. J Clin Microbiol 10 2: 144–6.

72. NielsenML, AlbertsenL, LettierG, NielsenJB, MortensenUH (2006) Efficient PCR-based gene targeting with a recyclable marker for Aspergillus nidulans. Fungal Genet Biol 43 1: 54–64.

73. OrejasM, EspesoEA, TilburnJ, SarkarS, ArstHNJr, et al. (1995) Activation of the Aspergillus PacC transcription factor in response to alkaline ambient pH requires proteolysis of the carboxy-terminal moiety. Genes Dev 9 13: 1622–32.

74. BradfordMM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72 248–54.

75. SchrettlM, BignellE, KraglC, JoechlC, RogersT, et al. (2004) Siderophore biosynthesis but not reductive iron assimilation is essential for Aspergillus fumigatus virulence. J Exp Med 200 9: 1213–9.

76. SaijoS, FujikadoN, FurutaT, ChungSH, KotakiH, et al. (2007) Dectin-1 is required for host defense against Pneumocystis carinii but not against Candida albicans. Nat Immunol 8 1: 39–46.

77. HuangDW, ShermanBT, LempickiRA (2008) Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat. Protocols 4 1: 44–57.

78. HuangDW, ShermanBT, LempickiRA (2009) Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic acids research 37 1: 1–13.

79. WalkerLA, MunroCA, de BruijnI, LenardonMD, McKinnonA, et al. (2008) Stimulation of chitin synthesis rescues Candida albicans from echinocandins. PLoS Pathog 4 4: e1000040.

80. Mora-MontesHM, NeteaMG, FerwerdaG, LenardonMD, BrownGD, et al. (2011) Recognition and blocking of innate immunity cells by Candida albicans chitin. Infect Immun 79 5: 1961–70.

81. PlaineA, WalkerL, Da CostaG, Mora-MontesHM, McKinnonA, et al. (2008) Functional analysis of Candida albicans GPI-anchored proteins: roles in cell wall integrity and caspofungin sensitivity. Fungal Genet Biol 45 10: 1404–14.

82. WheelerRT, FinkGR (2006) A drug-sensitive genetic network masks fungi from the immune system. PLoS Pathog 2 4: e35.

83. FujikawaT, KugaY, YanoS, YoshimiA, TachikiT, et al. (2009) Dynamics of cell wall components of Magnaporthe grisea during infectious structure development. Mol Microbiol 73 4: 553–70.

84. GravelatFN, EjzykowiczDE, ChiangLY, ChabotJC, UrbM, et al. (2010) Aspergillus fumigatus MedA governs adherence, host cell interactions and virulence. Cell Microbiol 12 4: 473–88.

85. TowbinH, StaehelinT, GordonJ (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A 76 9: 4350–4.