title: Lipid Flippase Subunit Cdc50 Mediates Drug Resistance and Virulence in Cryptococcus neoformans.
abstract: Cryptococcus neoformans is a human fungal pathogen and a major cause of fungal meningitis in immunocompromised individuals. Treatment options for cryptococcosis are limited. Of the two major antifungal drug classes, azoles are active against C. neoformans but exert a fungistatic effect, necessitating long treatment regimens and leaving open an avenue for emergence of azole resistance. Drugs of the echinocandin class, which target the glucan synthase and are fungicidal against a number of other fungal pathogens, such as Candida species, are ineffective against C. neoformans Despite the sensitivity of the target enzyme to the drug, the reasons for the innate resistance of C. neoformans to echinocandins remain unknown. To understand the mechanism of echinocandin resistance in C. neoformans, we screened gene disruption and gene deletion libraries for mutants sensitive to the echinocandin-class drug caspofungin and identified a mutation of CDC50, which encodes the β-subunit of membrane lipid flippase. We found that the Cdc50 protein localized to membranes and that its absence led to plasma membrane defects and enhanced caspofungin penetration into the cell, potentially explaining the increased caspofungin sensitivity. Loss of CDC50 also led to hypersensitivity to the azole-class drug fluconazole. Interestingly, in addition to functioning in drug resistance, CDC50 was also essential for fungal resistance to macrophage killing and for virulence in a murine model of cryptococcosis. Furthermore, the surface of cdc50Δ cells contained increased levels of phosphatidylserine, which has been proposed to act as a macrophage recognition signal. Together, these results reveal a previously unappreciated role of membrane lipid flippase in C. neoformans drug resistance and virulence.
  
  
  
  Cryptococcus neoformans is a fungal pathogen that is the most common cause of fungal meningitis, causing over 620,000 deaths annually. The treatment options for cryptococcosis are very limited. The most commonly used drugs are either fungistatic (azoles) or highly toxic (amphotericin B). Echinocandins are the newest fungicidal drug class that works well in treating candidiasis and aspergillosis, yet they are ineffective in treating cryptococcosis. In this study, we showed that the regulatory subunit of the lipid translocase (flippase), a protein that regulates the asymmetrical orientation of membrane lipids, is required for C. neoformans resistance to caspofungin, as well as for virulence during infection. This discovery identifies lipid flippase as a potential C. neoformans drug target, which plays an important role in the innate resistance of C. neoformans to echinocandins and in fungal virulence.
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Huang
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA School of Pharmaceutical Sciences, Southwest University, Chongqing, China.
authors.forename: Guojian
authors.initials: G
authors.lastname: Liao
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: Gregory M
authors.initials: GM
authors.lastname: Baker
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: Yina
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Lau
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: Padmaja
authors.initials: P
authors.lastname: Paderu
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA Department of Microbiology, Biochemistry and Molecular Genetics, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA.
authors.forename: David S
authors.initials: DS
authors.lastname: Perlin
authors.affiliation: Public Health Research Institute Center, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA Department of Microbiology, Biochemistry and Molecular Genetics, New Jersey Medical School, Rutgers University, Newark, New Jersey, USA xuech@njms.rutgers.edu.
authors.forename: Chaoyang
authors.initials: C
authors.lastname: Xue
mesh_terms: D000818:Animals; D000935:Antifungal Agents / Q000494:pharmacology*; D000077336:Caspofungin; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology / Q000473:pathology; D003453:Cryptococcosis / Q000382:microbiology; D003455:Cryptococcus neoformans / Q000187:drug effects / Q000201:enzymology / Q000235:genetics* / Q000472:pathogenicity*; D004195:Disease Models, Animal; D025141:Drug Resistance, Fungal / Q000235:genetics; D054714:Echinocandins / Q000494:pharmacology*; D015725:Fluconazole / Q000494:pharmacology; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D006801:Humans; D055666:Lipopeptides / Q000494:pharmacology*; D051379:Mice; D009154:Mutation; D010718:Phosphatidylserines / Q000276:immunology / Q000378:metabolism; D049294:Phospholipid Transfer Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014774:Virulence
chemical_list: D000935:Antifungal Agents; D054714:Echinocandins; D005656:Fungal Proteins; D055666:Lipopeptides; D010718:Phosphatidylserines; D049294:Phospholipid Transfer Proteins; D015725:Fluconazole; D000077336:Caspofungin