Browsing by Subject "Hyphae"

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    Characterization of the FKBP12-Encoding Genes in Aspergillus fumigatus.
    (PLoS One, 2015) Falloon, Katie; Juvvadi, Praveen R; Richards, Amber D; Vargas-Muñiz, José M; Renshaw, Hilary; Steinbach, William J
    Invasive aspergillosis, largely caused by Aspergillus fumigatus, is responsible for a growing number of deaths among immunosuppressed patients. Immunosuppressants such as FK506 (tacrolimus) that target calcineurin have shown promise for antifungal drug development. FK506-binding proteins (FKBPs) form a complex with calcineurin in the presence of FK506 (FKBP12-FK506) and inhibit calcineurin activity. Research on FKBPs in fungi is limited, and none of the FKBPs have been previously characterized in A. fumigatus. We identified four orthologous genes of FKBP12, the human FK506 binding partner, in A. fumigatus and designated them fkbp12-1, fkbp12-2, fkbp12-3, and fkbp12-4. Deletional analysis of the four genes revealed that the Δfkbp12-1 strain was resistant to FK506, indicating FKBP12-1 as the key mediator of FK506-binding to calcineurin. The endogenously expressed FKBP12-1-EGFP fusion protein localized to the cytoplasm and nuclei under normal growth conditions but also to the hyphal septa following FK506 treatment, revealing its interaction with calcineurin. The FKBP12-1-EGFP fusion protein didn't localize at the septa in the presence of FK506 in the cnaA deletion background, confirming its interaction with calcineurin. Testing of all deletion strains in the Galleria mellonella model of aspergillosis suggested that these proteins don't play an important role in virulence. While the Δfkbp12-2 and Δfkbp12-3 strains didn't show any discernable phenotype, the Δfkbp12-4 strain displayed slight growth defect under normal growth conditions and inhibition of the caspofungin-mediated "paradoxical growth effect" at higher concentrations of the antifungal caspofungin. Together, these results indicate that while only FKBP12-1 is the bona fide binding partner of FK506, leading to the inhibition of calcineurin in A. fumigatus, FKBP12-4 may play a role in basal growth and the caspofungin-mediated paradoxical growth response. Exploitation of differences between A. fumigatus FKBP12-1 and human FKBP12 will be critical for the generation of fungal-specific FK506 analogs to inhibit fungal calcineurin and treat invasive fungal disease.
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    Divergent Roles for cAMP-PKA Signaling in the Regulation of Filamentous Growth in Saccharomyces cerevisiae and Saccharomyces bayanus.
    (G3 (Bethesda, Md.), 2018-11) Kayikci, Ömur; Magwene, Paul M
    The cyclic AMP - Protein Kinase A (cAMP-PKA) pathway is an evolutionarily conserved eukaryotic signaling network that is essential for growth and development. In the fungi, cAMP-PKA signaling plays a critical role in regulating cellular physiology and morphological switches in response to nutrient availability. We undertook a comparative investigation of the role that cAMP-PKA signaling plays in the regulation of filamentous growth in two closely related budding yeast species, Saccharomyces cerevisiae and Saccharomyces bayanus Using chemical and genetic perturbations of this pathway and its downstream targets we discovered divergent roles for cAMP-PKA signaling in the regulation of filamentous growth. While cAMP-PKA signaling is required for the filamentous growth response in both species, increasing or decreasing the activity of this pathway leads to drastically different phenotypic outcomes. In S. cerevisiae, cAMP-PKA inhibition ameliorates the filamentous growth response while hyper-activation of the pathway leads to increased filamentous growth; the same perturbations in S. bayanus result in the obverse. Divergence in the regulation of filamentous growth between S. cerevisiae and S. bayanus extends to downstream targets of PKA, including several kinases, transcription factors, and effector proteins. Our findings highlight the potential for significant evolutionary divergence in gene network function, even when the constituent parts of such networks are well conserved.