Chad Rappleye
Research Interests
Our primary interests pertain to the molecular mechanisms that underlie the virulence of the respiratory fungal pathogen Histoplasma capsulatum, thecausative agent of histoplasmosis. In particular, our studies focus on the interaction betweenHistoplasma yeasts and their primary host cell, the mammalian macrophage. Unlike opportunistic pathogens, Histoplasma can cause disease even in immunocompetent hosts, indicating that Histoplasma has specific mechanisms designed to promote its pathogenesis. Our research has uncovered two of these mechanisms: masking of immunostimulatory cell wall molecules and production of extracellular defenses. Beta-glucan-containing fungal cell walls are potent activators of signaling receptors on the surface of macrophages. However, Histoplasma elaborates an alpha-linked glucan which masks beta-glucan polysaccharides from detection. We have also discovered a secreted endoglucanase which further removes any exposed beta-glucans on the cell wall, further enabling Histoplasma to be taken up by macrophages without alerting the macrophage. Phagocytosis by immune cells exposes fungi to antimicrobial reactive oxygen produced by phagocytes. Histoplasma yeast secrete a superoxide dismutase as well as a catalase which effectively destroy these phagocyte defense molecules, thereby enabling Histoplasma yeast to survive during infection.
To discover additional factors contributing to the pathogenesis of Histoplasma, our lab uses genomics-based analyses (transcriptomics and proteomics) as well as forward genetic screens. As candidate virulence genes are identified, their importance is assessed through the creation of loss-of-function strains. For this purpose we have developed and optimized molecular genetic techniques for Histoplasma including RNA inteference for gene product depletion as well as insertional mutagenesis using Agrobacterium tumefaciens T-DNA. Histoplasma is a natural pathogen of mammals and thus, virulence can be quantified with excellent animal models of respiratory and disseminated disease as well as in vitro culture systems which facilitate studies to dissect the effect of fungal proteins at the cellular level. To provide a more complete description of the molecular basis of fungal pathogenesis, our research also investigates the host side of the host-pathogen equation. Using retroviral and lentiviral RNA interference systems to reduce host gene functions, we can now test the involvement of host molecules in the response to Histoplasma infection. This ability to perform loss-of-function experiments in both the host and pathogen combine into a powerful system to define fungal virulence factors and the cognate host pathways targeted.
Our interests also extend to clinical application of fungal pathogen research, namely the development of improved fungal diagnostics and antifungal therapeutics. Our lab is part of a group of researchers at Ohio State pursuing antimicrobial drug discovery. We are screening multiple small molecule libraries and have discovered compounds that significantly inhibit fungal growth. One such compound has sub-micromolar in vitro efficacy against Histoplasma and Cryptococcus, two fungal pathogens that cause disease in otherwise healthy hosts and which are naturally resistant to the echinocandin-class antifungal drugs. In addition, we are applying our expertise in proteomics to discover new biomarkers that could provide specific and early indications of Histoplasma and Aspergillus infections. With improved diagnostics and antifungal options, we hope to reduce the morbidity and mortality resulting from systemic fungal infections.
Memberships
Relevant Publications
Beta-glucan Masking
Garfoot AL, Dearing KL, VanSchoiack AD, Wysocki VH, Rappleye CA. (2017) Eng1 and Exg8 Are the Major β-Glucanases Secreted by the Fungal Pathogen Histoplasma capsulatum. J Biol Chem 292:4801–4810. doi:10.1074/jbc.M116.762104 PMID: 28154008.
Garfoot AL, Shen Q, Wüthrich M, Klein BS, Rappleye CA. (2016) The Eng1 β-Glucanase Enhances Histoplasma Virulence by Reducing β-Glucan Exposure. mBio 7:e01388-01315. doi:10.1128/mBio.01388-15 PMID: 27094334.
Edwards JA, Alore EA, Rappleye CA. (2011) The yeast-phase virulence requirement for α-glucan synthase differs among Histoplasma capsulatum chemotypes. Eukaryot Cell 10:87–97. doi:10.1128/EC.00214-10 PMID: 21037179.
Rappleye CA, Eissenberg LG, Goldman WE. (2007) Histoplasma capsulatum alpha-(1,3)-glucan blocks innate immune recognition by the beta-glucan receptor. Proc Natl Acad Sci U S A 104:1366–1370. doi:10.1073/pnas.0609848104 PMID: 17227865.30.
Marion CL, Rappleye CA, Engle JT, Goldman WE. (2006) An alpha-(1,4)-amylase is essential for alpha-(1,3)-glucan production and virulence in Histoplasma capsulatum. Mol Microbiol 62:970–983. doi:10.1111/j.1365-2958.2006.05436.x PMID: 17038119.
Rappleye CA, Engle JT, Goldman WE. (2004) RNA interference in Histoplasma capsulatum demonstrates a role for alpha-(1,3)-glucan in virulence. Mol Microbiol 53:153–165. doi:10.1111/j.1365-2958.2004.04131.x PMID: 15225311.
ROS Defense
Holbrook ED, Smolnycki KA, Youseff BH, Rappleye CA. (2013) Redundant catalases detoxify phagocyte reactive oxygen and facilitate Histoplasma capsulatum pathogenesis. Infect Immun 81:2334–2346. doi:10.1128/IAI.00173-13 PMID: 23589579.
Youseff BH, Holbrook ED, Smolnycki KA, Rappleye CA. (2012) Extracellular superoxide dismutase protects Histoplasma yeast cells from host-derived oxidative stress. PLoS Pathog 8:e1002713. doi:10.1371/journal.ppat.1002713 PMID: 22615571.
Intracellular Pathogenesis Mechanisms
Brechting PJ, Shah C, Rakotondraibe L, Shen Q, Rappleye CA. (2023) Histoplasma capsulatum requires peroxisomes for multiple virulence functions including siderophore biosynthesis. mBio 14(4):e0328422. doi: 10.1128/mbio.03284-22. PMID: 37432032
Shen Q, Gonzalez-Mireles A, Ray SC, Rappleye CA. (2023) Histoplasma capsulatum Relies on Tryptophan Biosynthesis To Proliferate within the Macrophage Phagosome. Infect Immun 91(6):e0005923. doi: 10.1128/iai.00059-23. PMID: 37184383
Ray SC, Rappleye CA. (2022) Mac1-Dependent Copper Sensing Promotes Histoplasma Adaptation to the Phagosome during Adaptive Immunity. mBio 13:e0377321. doi:10.1128/mbio.03773-21 PMID: 35404120.
Shen Q, Ray SC, Evans HM, Deepe GS, Rappleye CA. (2020) Metabolism of Gluconeogenic Substrates by an Intracellular Fungal Pathogen Circumvents Nutritional Limitations within Macrophages. mBio 11:e02712-19. doi:10.1128/mBio.02712-19 PMID: 32265333.
Shen Q, Beucler MJ, Ray SC, Rappleye CA. (2018) Macrophage activation by IFN-γ triggers restriction of phagosomal copper from intracellular pathogens. PLoS Pathog 14:e1007444. doi:10.1371/journal.ppat.1007444 PMID: 30452484.
Garfoot AL, Goughenour KD, Wüthrich M, Rajaram MVS, Schlesinger LS, Klein BS, Rappleye CA. (2018) O-Mannosylation of Proteins Enables Histoplasma Yeast Survival at Mammalian Body Temperatures. mBio 9:e02121-17. doi:10.1128/mBio.02121-17 PMID: 29295913.
Garfoot AL, Zemska O, Rappleye CA. (2014) Histoplasma capsulatum depends on de novo vitamin biosynthesis for intraphagosomal proliferation. Infect Immun 82:393–404. doi:10.1128/IAI.00824-13 PMID: 24191299.
Genomics/Transcriptomics/Proteomics
Goughenour KD, Whalin J, Slot JC, Rappleye CA. (2021) Diversification of Fungal Chitinases and Their Functional Differentiation in Histoplasma capsulatum. Mol Biol Evol 38:1339–1355. doi:10.1093/molbev/msaa293 PMID: 33185664.
Longo LVG, Ray SC, Puccia R, Rappleye CA. (2018) Characterization of the APSES-family transcriptional regulators of Histoplasma capsulatum. FEMS Yeast Res 18:foy087. doi:10.1093/femsyr/foy087 PMID: 30101348.
Edwards JA, Chen C, Kemski MM, Hu J, Mitchell TK, Rappleye CA. (2013) Histoplasma yeast and mycelial transcriptomes reveal pathogenic-phase and lineage-specific gene expression profiles. BMC Genomics 14:695. doi:10.1186/1471-2164-14-695 PMID: 24112604.
Holbrook ED, Edwards JA, Youseff BH, Rappleye CA. (2011) Definition of the extracellular proteome of pathogenic-phase Histoplasma capsulatum. J Proteome Res 10:1929–1943. doi:10.1021/pr1011697 PMID: 21291285.
Molecular Genetic Tools
Rappleye CA. (2023) Targeted gene deletions in the dimorphic fungal pathogen Histoplasma using an optimized episomal CRISPR/Cas9 system. mSphere 8(4):e0017823. doi: 10.1128/msphere.00178-23. PMID: 37389430
Kemski MM, Stevens B, Rappleye CA. (2013) Spectrum of T-DNA integrations for insertional mutagenesis of Histoplasma capsulatum. Fungal Biol 117:41–51. doi:10.1016/j.funbio.2012.11.004 PMID: 23332832.
Youseff BH, Rappleye CA. (2012) RNAi-based gene silencing using a GFP sentinel system in Histoplasma capsulatum. Methods Mol Biol 845:151–164. doi:10.1007/978-1-61779-539-8_10 PMID: 22328373.
Zemska O, Rappleye CA. (2012) Agrobacterium-mediated insertional mutagenesis in Histoplasma capsulatum. Methods Mol Biol 845:51–66. doi:10.1007/978-1-61779-539-8_4 PMID: 22328367.Edwards JA, Zemska O, Rappleye CA. (2011) Discovery of a role for Hsp82 in Histoplasma virulence through a quantitative screen for macrophage lethality. Infect Immun 79:3348–3357. doi:10.1128/IAI.05124-11 PMID: 21606189.
Youseff BH, Dougherty JA, Rappleye CA. (2009) Reverse genetics through random mutagenesis in Histoplasma capsulatum. BMC Microbiol 9:236. doi:10.1186/1471-2180-9-236 PMID: 19919692.
Antifungals and Diagnostics
Ishita K, Stefanopoulos S, Khalil A, Cheng X, Tjarks W, Rappleye CA. (2018) Synthesis and biological evaluation of aminothiazoles against Histoplasma capsulatum and Cryptococcus neoformans. Bioorg Med Chem 26:2251–2261. doi:10.1016/j.bmc.2018.01.024 PMID: 29580849.
Goughenour KD, Rappleye CA. (2017) Antifungal therapeutics for dimorphic fungal pathogens. Virulence 8:211–221. doi:10.1080/21505594.2016.1235653 PMID: 27646561.
Khalil A, Edwards JA, Rappleye CA, Tjarks W. (2015) Design, synthesis, and biological evaluation of aminothiazole derivatives against the fungal pathogens Histoplasma capsulatum and Cryptococcus neoformans. Bioorg Med Chem 23:532–547. doi:10.1016/j.bmc.2014.12.006 PMID: 25543205.
Goughenour KD, Balada-Llasat J-M, Rappleye CA. (2015) Quantitative Microplate-Based Growth Assay for Determination of Antifungal Susceptibility of Histoplasma capsulatum Yeasts. J Clin Microbiol 53:3286–3295. doi:10.1128/JCM.00795-15 PMID: 26246483.
Holbrook ED, Kemski MM, Richer SM, Wheat LJ, Rappleye CA. (2014) Glycosylation and immunoreactivity of the Histoplasma capsulatum Cfp4 yeast-phase exoantigen. Infect Immun 82:4414–4425. doi:10.1128/IAI.01893-14 PMID: 25114108.
Edwards JA, Kemski MM, Rappleye CA. (2013) Identification of an aminothiazole with antifungal activity against intracellular Histoplasma capsulatum. Antimicrob Agents Chemother 57:4349–4359. doi:10.1128/AAC.00459-13 PMID: 23817367.
Fungal Pathogenesis Reviews
Brechting PJ, Rappleye CA. (2019) Histoplasma Responses to Nutritional Immunity Imposed by Macrophage Activation. J Fungi (Basel) 5:45. doi:10.3390/jof5020045 PMID: 31195617.
Ray SC, Rappleye CA. (2018) Flying under the radar: Histoplasma capsulatum avoidance of innate immune recognition. Semin Cell Dev Biol https://doi.org/10.1016/j.semcdb.2018.03.009 doi:10.1016/j.semcdb.2018.03.009 PMID: 29551572.
Shen Q, Rappleye CA. (2017) Differentiation of the fungus Histoplasma capsulatum into a pathogen of phagocytes. Curr Opin Microbiol 40:1–7. doi:10.1016/j.mib.2017.10.003 PMID: 29096192.
Garfoot AL, Rappleye CA. (2016) Histoplasma capsulatum surmounts obstacles to intracellular pathogenesis. FEBS J 283:619–633. doi:10.1111/febs.13389 PMID: 26235362.
Edwards JA, Rappleye CA. (2011) Histoplasma mechanisms of pathogenesis--one portfolio doesn’t fit all. FEMS Microbiol Lett 324:1–9. doi:10.1111/j.1574-6968.2011.02363.x PMID: 22092757.
Holbrook ED, Rappleye CA. (2008) Histoplasma capsulatum pathogenesis: making a lifestyle switch. Curr Opin Microbiol 11:318–324. doi:10.1016/j.mib.2008.05.010 PMID: 18573684.
Rappleye CA. (2014) Molecular Mechanisms of Histoplasma Pathogenesis, p. 129–140. In Kurzai O (ed.), Human Fungal Pathogens, 2nd ed. Springer Press, New York
Rappleye CA, Goldman WE. (2008) Fungal stealth technology. Trends Immunol 29:18–24. doi:10.1016/j.it.2007.10.001 PMID: 18054285.
Rappleye CA, Goldman WE. (2006) Defining virulence genes in the dimorphic fungi. Annu Rev Microbiol 60:281–303. doi:10.1146/annurev.micro.59.030804.121055 PMID: 16753032.