Titan cells formation in Cryptococcus neoformans is finely tuned by environmental conditions and modulated by positive and negative genetic regulators
| Autor: | James A. Fraser, Liliane Mukaremera, John R. Perfect, Arturo Casadevall, Françoise Dromer, Benjamin Hommel, Christopher A. Desjardins, Alexandre Alanio, Aude Sturny-Leclère, Guilhem Janbon, Christina A. Cuomo, Carolina Coelho, Kirsten Nielsen, Radames J. B. Cordero |
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| Přispěvatelé: | Mycologie moléculaire - Molecular Mycology, Centre National de la Recherche Scientifique (CNRS)-Institut Pasteur [Paris], Université Paris Diderot - Paris 7 (UPD7), Laboratoire de Parasitologie-Mycologie [CHU Saint Louis, Paris], Groupe Hospitalier Saint Louis - Lariboisière - Fernand Widal [Paris], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), University of Minnesota [Twin Cities] (UMN), University of Minnesota System, Johns Hopkins University School of Medicine [Baltimore], Broad Institute of MIT and Harvard (BROAD INSTITUTE), Harvard Medical School [Boston] (HMS)-Massachusetts Institute of Technology (MIT)-Massachusetts General Hospital [Boston], Biologie des ARN des Pathogènes fongiques - RNA Biology of Fungal Pathogens, Institut Pasteur [Paris], Duke University [Durham], University of Queensland [Brisbane], BH’s salary was funded by Assistance Publique-Hôpitaux de Paris and Institut Pasteur (Poste d’ accueil APHP/CNRS/Institut Pasteur). http://recherche.aphp.fr/candidatures-internes/ KN grant funding National Institutes of Health, National Institute of Allergy and Infectious Diseases (NIAID) grant R01AI080275. https://www.niaid.nih.gov AC is supported in part by 5R01HL059842, 5R01AI033774, 5R37AI033142, and 5R01AI052733. https://www.niaid.nih.gov CAD and CAC were supported by National Institute of Allergy and Infectious Diseases, National Institutes of Health, Department of Health and Human Services grant number U19AI110818. https://www.niaid.nih.gov The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript., Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris] (IP), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS) |
| Jazyk: | angličtina |
| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Cell division Cell Vacuole MESH: Quorum Sensing MESH: Lung Diseases Fungal MESH: Animals Biology (General) [SDV.MP.MYC]Life Sciences [q-bio]/Microbiology and Parasitology/Mycology 2. Zero hunger education.field_of_study 0303 health sciences Chemistry MESH: Cryptococcus neoformans Phenotype Cell biology medicine.anatomical_structure Interaction with host symbols Titan (rocket family) MESH: Mutation QH301-705.5 030106 microbiology Immunology Population MESH: Cryptococcosis Biology MESH: Phenotype Microbiology Cell wall symbols.namesake 03 medical and health sciences MESH: Hyphae MESH: Mice Inbred C57BL Virology Genetics medicine Progenitor cell education Molecular Biology MESH: Mice 030304 developmental biology Cryptococcus neoformans MESH: Humans 030306 microbiology fungi MESH: Host-Pathogen Interactions MESH: Models Biological RC581-607 biology.organism_classification In vitro Nucleic acid biosynthesis Parasitology Immunologic diseases. Allergy MESH: Disease Models Animal MESH: Genes Fungal Biogenesis |
| Zdroj: | PLoS Pathogens PLoS Pathogens, Public Library of Science, 2018, 14 (5), pp.e1006982. ⟨10.1371/journal.ppat.1006982⟩ PLoS Pathogens, 2018, 14 (5), pp.e1006982. ⟨10.1371/journal.ppat.1006982⟩ PLoS Pathogens, Vol 14, Iss 5, p e1006982 (2018) |
| ISSN: | 1553-7366 1553-7374 |
| DOI: | 10.1371/journal.ppat.1006982⟩ |
| Popis: | A remarkable aspect of the human fungal pathogen Cryptococcus neoformans is the morphological changes triggered by the interaction with the host. During infection, a specific morphological change in cell size is observed, particularly in lung tissue, from regular 5 to 7 µm cells (RCs) to titan cells (TCs, > 10 µm and up to 100 µm). However, the stable and reproducible generation of large quantity of TCs was only possible during experimental infection. We implemented here in vitro conditions allowing TCs generation with the aim to understand the ancestry of TC, the environmental determinants of TCs formation and perform easily phenotype/genotype analysis to uncover genetic factors underlying TCs formation. This paper reports robust in vitro conditions that generates yeasts cells harboring the main characteristics of TCs: large (>10 µm), uninucleate and polyploid cells harboring a single large vacuole with a dense capsule surrounding a thickened melanized cell wall. We observed that TCs formation begins as soon as 8 hours after induction, with a maximal proportion of TCs obtained after 24 hours. TCs derived for the initial oldest cells (mother cells) rather than from daughter cells. Hypoxia, nutrient starvation and low pH stresses were the main factors that trigger TCs formation, while quorum sensing factors like the initial inoculum concentration and the addition of pantothenic acid also impacted TCs formation. Antifungal drugs, resulting in inhibition of ergosterol (fluconazole), or proteins and nucleic acids (flucytosine), altered TCs formation, as did hosts-related products such as serum, phospholipids and anti-capsular antibodies in our settings. We then explored genetic factors important for TCs formation using two approaches. Using strains from H99 lineage strains among which few genetic differences have been described, we showed that TCs formation was dependent on Lmp1, Sgf29 and Gpr4/5-Rim101 proteins. Then, based on a the analysis of natural Pkr1 loss-of-function clinical isolates and serial clinical isolates, we observed the important role of Pkr1 protein, a negative regulator of the cyclic adenosine monophosphate / protein kinase A (cAMP/PKA), for TCs formation. These strains also emphasize the structural and functional relationship between Pkr1 and Pka. Our results provide new insights into TCs biogenesis with identification of multiple important factors/pathways involved. The implementation of such standardized and robust in vitro conditions pave the way for future research focusing on the genetic basis of TCs biogenesis, biology of TCs and the ontology of morphological changes in Cryptococcus neoformans. Author Summary Cryptococcus neoformans is a yeast that is capable of morphological change upon interaction with host. Particularly, in the lung of infected mice, a subpopulation of yeast evolves toward gigantism (titan cells size from 10 to 100 µm) that include other characteristics such as thickened cell wall, dense capsule, polyploidization, large vacuole with peripheral nucleus and cellular organelles. The generation of large number of such cells outside the lung of mice have been described but was not reproducible nor standardized. We here report standardized, reproducible, robust conditions of generation of titan cells and explored the environmental and genetic factors underlying the genesis of these cells. We showed that Titan cells were generated upon stresses such as change in the medium, nutrient deprivation, hypoxia and low pH. Using collection of well characterized reference strains and clinical isolates, we validated with our model the AMPc/PKA/Rim pathway as the major genetic determinant of titan cell formation. This study opens the way for a more comprehensive picture of the ontology of morphological changes in Cryptococcus neoformans. |
| Databáze: | OpenAIRE |
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