Role of Calcineurin in Fungal Virulence

Summary

Principal Investigator: JOSEPH B HEITMAN
Abstract: The incidence of fungal infection is increasing and yet available antifungal drugs are limited, some are toxic, and drug resistant strains are emerging. We have elucidated a conserved signal transduction cascade that controls virulence of Cryptococcus neoformans, the leading cause of fungal meningitis. The central element of this virulence pathway is the calcium-calmodulin activated protein phosphatase calcineurin, which is the molecular target of the immunosuppressive antifungal drugs cyclosporin A and FK506. C. neoformans mutants lacking either the catalytic A or the regulatory B subunit of calcineurin are inviable at 37 degrees C and other stress conditions and, as a consequence, are avirulent in animal models. In studies supported by this award, we identified: 1) the calcineurin B regulatory subunit and calmodulin, 2) the calcineurin binding protein (Cbpl) that is a conserved regulator or effector and which is the founding member of a protein family conserved from fungi to humans, and 3) the novel C2 domain protein Cts1 that may function as a downstream effector of the calcineurin signaling pathway to promote cell wall biogenesis and growth at 37degrees C. In parallel we discovered that calcineurin is required for virulence of Candida albicans, the most common human fungal pathogen. C. albicans cnb1/cnb1 mutants lacking the calcineurin B regulatory subunit are severely attenuated in animal models. Yet, in contrast to C. neoformans calcineurin is not required for growth of C. albicans at 37 degrees C. Instead, calcineurin is necessary for C. albicans to survive and proliferate in serum. These studies illustrate how a conserved signaling cascade has been co-opted to control virulence of two divergent fungal pathogens by unique molecular mechanisms. Here we propose to delineate this molecular virulence cascade in both C. neoformans and C. albicans. Importantly, this pathway can be targeted for therapeutic intervention using non-immunosuppressive calcineurin inhibitors that retain antifungal activity and synergistic drug combinations that we have discovered.
Funding Period: 2009-07-14 - 2009-10-31
more information: NIH RePORT

Top Publications

  1. pmc Surfactant protein D facilitates Cryptococcus neoformans infection
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina, USA
    Infect Immun 80:2444-53. 2012
  2. pmc The C2 domain protein Cts1 functions in the calcineurin signaling circuit during high-temperature stress responses in Cryptococcus neoformans
    Eanas F Aboobakar
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:1714-23. 2011
  3. pmc Association of calcineurin with the COPI protein Sec28 and the COPII protein Sec13 revealed by quantitative proteomics
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
    PLoS ONE 6:e25280. 2011
  4. pmc Calcineurin colocalizes with P-bodies and stress granules during thermal stress in Cryptococcus neoformans
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:1396-402. 2011
  5. pmc Calcineurin controls drug tolerance, hyphal growth, and virulence in Candida dubliniensis
    Ying Lien Chen
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:803-19. 2011
  6. pmc Surfactant protein D binding to Aspergillus fumigatus hyphae is calcineurin-sensitive
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Med Mycol 48:580-8. 2010
  7. pmc Septins enforce morphogenetic events during sexual reproduction and contribute to virulence of Cryptococcus neoformans
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Mol Microbiol 75:658-75. 2010
  8. pmc Elucidating the Candida albicans calcineurin signaling cascade controlling stress response and virulence
    Jennifer L Reedy
    Department of Molecular Genetics and Microbiology, Duke University, Box 3546 Research Drive, 322 CARL Bldg, Durham, NC 27710, USA
    Fungal Genet Biol 47:107-16. 2010
  9. pmc Surfactant protein D increases phagocytosis of hypocapsular Cryptococcus neoformans by murine macrophages and enhances fungal survival
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Infect Immun 77:2783-94. 2009
  10. pmc Identification of ENA1 as a virulence gene of the human pathogenic fungus Cryptococcus neoformans through signature-tagged insertional mutagenesis
    Alexander Idnurm
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Eukaryot Cell 8:315-26. 2009

Scientific Experts

  • Lukasz Kozubowski
  • W J Steinbach
  • Natalie A Afshari
  • Alexander Idnurm
  • Joseph Heitman
  • Scarlett Geunes-Boyer
  • Jennifer L Reedy
  • Jo Rae Wright
  • Ying Lien Chen
  • Eanas F Aboobakar
  • John R Perfect
  • Scott G Filler
  • Priya Uppuluri
  • Weihua Fan
  • Chiatogu Onyewu
  • Kaihei Kojima
  • Felicia J Walton
  • Michael F Beers
  • Xuying Wang
  • Anna Averette
  • David R Andes
  • Emma L Morrison
  • Ursela G Bigol
  • Fedelino F Malbas
  • Fitz Gerald S Silao
  • Jeniel E Nett
  • Norma V Solis
  • Alexandra Brand
  • Timothy N Oliver
  • Jennifer K Lodge
  • Guilhem Janbon
  • David Andes
  • Jeniel Nett
  • Julia Breger
  • Eleftherios Mylonakis
  • Shahid Husain
  • Timothy L Pruett
  • Yong Sun Bahn
  • Michael Ison
  • Nina Singh

Detail Information

Publications17

  1. pmc Surfactant protein D facilitates Cryptococcus neoformans infection
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina, USA
    Infect Immun 80:2444-53. 2012
    ..This study establishes a new paradigm for the role played by SP-D during host responses to C. neoformans and consequently imparts insight into potential future preventive and/or treatment strategies for cryptococcosis...
  2. pmc The C2 domain protein Cts1 functions in the calcineurin signaling circuit during high-temperature stress responses in Cryptococcus neoformans
    Eanas F Aboobakar
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:1714-23. 2011
    ..Taken together, these findings support potential roles for Cts1 in endocytic trafficking, mRNA processing, and cytokinesis and suggest that Cts1 is a substrate of calcineurin during high-temperature stress responses...
  3. pmc Association of calcineurin with the COPI protein Sec28 and the COPII protein Sec13 revealed by quantitative proteomics
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina, United States of America
    PLoS ONE 6:e25280. 2011
    ..Cna1 exhibited a dramatic change in subcellular localization during high temperature stress from diffuse cytoplasmic to ER-associated puncta and the mother-bud neck and co-localized with Sec28 and Sec13...
  4. pmc Calcineurin colocalizes with P-bodies and stress granules during thermal stress in Cryptococcus neoformans
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:1396-402. 2011
    ..These results support a model in which calcineurin orchestrates thermal stress responses by associating with sites of mRNA processing...
  5. pmc Calcineurin controls drug tolerance, hyphal growth, and virulence in Candida dubliniensis
    Ying Lien Chen
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Eukaryot Cell 10:803-19. 2011
    ..That calcineurin is required for drug tolerance and virulence makes fungus-specific calcineurin inhibitors attractive candidates for combination therapy with azoles or echinocandins against emerging C. dubliniensis infections...
  6. pmc Surfactant protein D binding to Aspergillus fumigatus hyphae is calcineurin-sensitive
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Med Mycol 48:580-8. 2010
    ..Elucidation of the interaction between lung innate immune factors and A. fumigatus could lead to the development of novel therapeutic interventions...
  7. pmc Septins enforce morphogenetic events during sexual reproduction and contribute to virulence of Cryptococcus neoformans
    Lukasz Kozubowski
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Mol Microbiol 75:658-75. 2010
    ..Strains lacking CDC3 or CDC12 exhibit significantly reduced virulence in a Galleria mellonella model of infection. Thus, C. neoformans septins are vital to morphology of the hyphae and contribute to virulence...
  8. pmc Elucidating the Candida albicans calcineurin signaling cascade controlling stress response and virulence
    Jennifer L Reedy
    Department of Molecular Genetics and Microbiology, Duke University, Box 3546 Research Drive, 322 CARL Bldg, Durham, NC 27710, USA
    Fungal Genet Biol 47:107-16. 2010
    ..These studies extend our understanding of the C. albicans calcineurin signaling cascade and its host-niche specific role in virulence...
  9. pmc Surfactant protein D increases phagocytosis of hypocapsular Cryptococcus neoformans by murine macrophages and enhances fungal survival
    Scarlett Geunes-Boyer
    Department of Cell Biology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Infect Immun 77:2783-94. 2009
    ..These findings provide evidence that C. neoformans subverts host defense mechanisms involving surfactant, establishing a novel virulence paradigm that may be targeted for therapy...
  10. pmc Identification of ENA1 as a virulence gene of the human pathogenic fungus Cryptococcus neoformans through signature-tagged insertional mutagenesis
    Alexander Idnurm
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Eukaryot Cell 8:315-26. 2009
    ..Signature-tagged mutagenesis is an effective strategy for the discovery of new virulence genes in fungal pathogens of animals...
  11. pmc Synergistic effect of calcineurin inhibitors and fluconazole against Candida albicans biofilms
    Priya Uppuluri
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, North Carolina 27710, USA
    Antimicrob Agents Chemother 52:1127-32. 2008
    ..These findings reveal that calcineurin contributes to fluconazole resistance of biofilms and provide evidence that synergistic drug combinations may prove efficacious as novel therapeutic interventions to treat or prevent biofilms...
  12. pmc Calcineurin inhibition or mutation enhances cell wall inhibitors against Aspergillus fumigatus
    William J Steinbach
    Division of Pediatric Infectious Diseases, Duke University Medical Center, Durham, NC 27710, USA
    Antimicrob Agents Chemother 51:2979-81. 2007
    ..Quantification of 1,3-beta-d-glucan revealed decreased amounts in the calcineurin A (DeltacnaA) mutant. Calcineurin can be an excellent adjunct therapeutic target in combination with other cell wall inhibitors against A. fumigatus...
  13. pmc Eca1, a sarcoplasmic/endoplasmic reticulum Ca2+-ATPase, is involved in stress tolerance and virulence in Cryptococcus neoformans
    Weihua Fan
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Infect Immun 75:3394-405. 2007
    ..Eca1 is likely involved in maintaining ER function, thus contributing to stress tolerance and virulence acting in parallel with Ca2+-calcineurin signaling...
  14. pmc Calcineurin promotes infection of the cornea by Candida albicans and can be targeted to enhance fluconazole therapy
    Chiatogu Onyewu
    Department of Ophthalmology, Wadsworth Bldg, Box 3802, Erwin Road, Duke University Eye Center, Durham, NC 27710, USA
    Antimicrob Agents Chemother 50:3963-5. 2006
    ..albicans infection more rapidly than monotherapy with either drug. Calcineurin, the target of CsA, was also found to contribute to pathogenicity...
  15. pmc Conserved elements of the RAM signaling pathway establish cell polarity in the basidiomycete Cryptococcus neoformans in a divergent fashion from other fungi
    Felicia J Walton
    Department of Molecular Genetics and Microbiology, Duke University Medical Center, Durham, NC 27710, USA
    Mol Biol Cell 17:3768-80. 2006
    ..neoformans but not in S. cerevisiae. These results indicate that conserved signaling pathways serve both similar and divergent cellular roles in morphogenesis in these divergent organisms...
  16. pmc Immunotherapy with tacrolimus (FK506) does not select for resistance to calcineurin inhibitors in Candida albicans isolates from liver transplant patients
    Jennifer L Reedy
    Department of Molecular Genetics and Microbiology, 322 CARL Building, Box 3546, Duke University Medical Center, Research Drive, Durham, NC 27710, USA
    Antimicrob Agents Chemother 50:1573-7. 2006
    ..We were unable to detect a difference in susceptibility to calcineurin inhibitors in combination with fluconazole, serum, or calcium in these isolates...
  17. ncbi Calcineurin, Mpk1 and Hog1 MAPK pathways independently control fludioxonil antifungal sensitivity in Cryptococcus neoformans
    Kaihei Kojima
    Department of Molecular Genetics and Microbiology, Durham, NC 27710, USA
    Microbiology 152:591-604. 2006
    ..These studies provide evidence that the broad-spectrum antifungal drug fludioxonil exerts its action via activation of the Hog1 MAPK pathway and provide insight into novel targets for synergistic antifungal drug combinations...