Calcineurin regulatory subunit is essential for virulence and mediates interactions with FKBP12-FK506 in Cryptococcus neoformans.
Calcineurin is a Ca2+-calmodulin-regulated protein phosphatase that is the target of the immunosuppressive drugs cyclosporin A and FK506. Calcineurin is a heterodimer composed of a catalytic A and a regulatory B subunit. In previous studies, the calcineurin A homologue was identified and shown to be required for growth at 37 degrees C and hence for virulence of the pathogenic fungus Cryptococcus neoformans. Here, we identify the gene encoding the calcineurin B regulatory subunit and demonstrate that calcineurin B is also required for growth at elevated temperature and virulence. We show that the FKR1-1 mutation, which confers dominant FK506 resistance, results from a 6 bp duplication generating a two-amino-acid insertion in the latch region of calcineurin B. This mutation was found to reduce FKBP12-FK506 binding to calcineurin both in vivo and in vitro. Molecular modelling based on the FKBP12-FK506-calcineurin crystal structure illustrates how this mutation perturbs drug interactions with the phosphatase target. In summary, our studies reveal a central role for calcineurin B in virulence and antifungal drug action in the human fungal pathogen C. neoformans.
Department of Genetics, 322 CARL Bldg., Box 3546, Duke University Medical Center, Research Drive, Durham, NC 27710, USA.
D S
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Fox
M C
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Cruz
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Sia
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Ke
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Cox
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Cardenas
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Heitman
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019703:Calcineurin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003453:Cryptococcosis / Q000382:microbiology; D003455:Cryptococcus neoformans / Q000235:genetics / Q000254:growth & development / Q000378:metabolism* / Q000472:pathogenicity; D004271:DNA, Fungal; D004195:Disease Models, Animal; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D006361:Heating; D006801:Humans; D051379:Mice; D008811:Mice, Inbred DBA; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017433:Protein Structure, Secondary; D011995:Recombination, Genetic; D017386:Sequence Homology, Amino Acid; D016559:Tacrolimus / Q000737:chemistry / Q000378:metabolism*; D022061:Tacrolimus Binding Protein 1A / Q000737:chemistry / Q000378:metabolism*; D014774:Virulence
D004271:DNA, Fungal; D005656:Fungal Proteins; D019703:Calcineurin; D022061:Tacrolimus Binding Protein 1A; D016559:Tacrolimus