A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>

ABSTRACT Mucormycosis is an emerging lethal fungal infection in immunocompromised patients. Mucor circinelloides is a causal agent of mucormycosis and serves as a model system to understand genetics in Mucorales. Calcineurin is a conserved virulence factor in many pathogenic fungi, and calcineurin i...

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Autores principales:	Sandeep Vellanki, R. Blake Billmyre, Alejandra Lorenzen, Micaela Campbell, Broderick Turner, Eun Young Huh, Joseph Heitman, Soo Chan Lee
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	Mucor mucormycosis amino acid permease calcineurin dimorphism drug resistance mechanisms Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/12849759825f4cdd96318e89c59485e8
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spelling	oai:doaj.org-article:12849759825f4cdd96318e89c59485e82021-11-15T15:56:58ZA Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>10.1128/mBio.02949-192150-7511https://doaj.org/article/12849759825f4cdd96318e89c59485e82020-02-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02949-19https://doaj.org/toc/2150-7511ABSTRACT Mucormycosis is an emerging lethal fungal infection in immunocompromised patients. Mucor circinelloides is a causal agent of mucormycosis and serves as a model system to understand genetics in Mucorales. Calcineurin is a conserved virulence factor in many pathogenic fungi, and calcineurin inhibition or deletion of the calcineurin regulatory subunit (CnbR) in Mucor results in a shift from hyphal to yeast growth. We analyzed 36 calcineurin inhibitor-resistant or bypass mutants that exhibited hyphal growth in the presence of calcineurin inhibitors or in the yeast-locked cnbRΔ mutant background without carrying any mutations in known calcineurin components. We found that a majority of the mutants had altered sequence in a gene, named here bycA (bypass of calcineurin). bycA encodes an amino acid permease. We verified that both the bycAΔ single mutant and the bycAΔ cnbRΔ double mutant are resistant to calcineurin inhibitor FK506, thereby demonstrating a novel mechanism of resistance against calcineurin inhibitors. We also found that the level of expression of bycA was significantly higher in the wild-type strain treated with FK506 and in the cnbRΔ mutants but was significantly lower in the wild-type strain without FK506 treatment. These findings suggest that bycA is a negative regulator of hyphal growth and/or a positive regulator of yeast growth in Mucor and that calcineurin suppresses expression of the bycA gene at the mRNA level to promote hyphal growth. BycA is involved in the Mucor hypha-yeast transition as our data demonstrate positive correlations among bycA expression, protein kinase A activity, and Mucor yeast growth. Also, calcineurin, independently of its role in morphogenesis, contributes to virulence traits, including phagosome maturation blockade, host cell damages, and proangiogenic growth factor induction during interactions with hosts. IMPORTANCE Mucor is intrinsically resistant to most known antifungals, which makes mucormycosis treatment challenging. Calcineurin is a serine/threonine phosphatase that is widely conserved across eukaryotes. When calcineurin function is inhibited in Mucor, growth shifts to a less virulent yeast growth form, which makes calcineurin an attractive target for development of new antifungal drugs. Previously, we identified two distinct mechanisms through which Mucor can become resistant to calcineurin inhibitors involving Mendelian mutations in the gene for FKBP12, including mechanisms corresponding to calcineurin A or B subunits and epimutations silencing the FKBP12 gene. Here, we identified a third novel mechanism where loss-of-function mutations in the amino acid permease corresponding to the bycA gene contribute to resistance against calcineurin inhibitors. When calcineurin activity is absent, BycA can activate protein kinase A (PKA) to promote yeast growth via a cAMP-independent pathway. Our data also show that calcineurin activity contributes to host-pathogen interactions primarily in the pathogenesis of Mucor.Sandeep VellankiR. Blake BillmyreAlejandra LorenzenMicaela CampbellBroderick TurnerEun Young HuhJoseph HeitmanSoo Chan LeeAmerican Society for MicrobiologyarticleMucormucormycosisamino acid permeasecalcineurindimorphismdrug resistance mechanismsMicrobiologyQR1-502ENmBio, Vol 11, Iss 1 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Mucor mucormycosis amino acid permease calcineurin dimorphism drug resistance mechanisms Microbiology QR1-502
spellingShingle	Mucor mucormycosis amino acid permease calcineurin dimorphism drug resistance mechanisms Microbiology QR1-502 Sandeep Vellanki R. Blake Billmyre Alejandra Lorenzen Micaela Campbell Broderick Turner Eun Young Huh Joseph Heitman Soo Chan Lee A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
description	ABSTRACT Mucormycosis is an emerging lethal fungal infection in immunocompromised patients. Mucor circinelloides is a causal agent of mucormycosis and serves as a model system to understand genetics in Mucorales. Calcineurin is a conserved virulence factor in many pathogenic fungi, and calcineurin inhibition or deletion of the calcineurin regulatory subunit (CnbR) in Mucor results in a shift from hyphal to yeast growth. We analyzed 36 calcineurin inhibitor-resistant or bypass mutants that exhibited hyphal growth in the presence of calcineurin inhibitors or in the yeast-locked cnbRΔ mutant background without carrying any mutations in known calcineurin components. We found that a majority of the mutants had altered sequence in a gene, named here bycA (bypass of calcineurin). bycA encodes an amino acid permease. We verified that both the bycAΔ single mutant and the bycAΔ cnbRΔ double mutant are resistant to calcineurin inhibitor FK506, thereby demonstrating a novel mechanism of resistance against calcineurin inhibitors. We also found that the level of expression of bycA was significantly higher in the wild-type strain treated with FK506 and in the cnbRΔ mutants but was significantly lower in the wild-type strain without FK506 treatment. These findings suggest that bycA is a negative regulator of hyphal growth and/or a positive regulator of yeast growth in Mucor and that calcineurin suppresses expression of the bycA gene at the mRNA level to promote hyphal growth. BycA is involved in the Mucor hypha-yeast transition as our data demonstrate positive correlations among bycA expression, protein kinase A activity, and Mucor yeast growth. Also, calcineurin, independently of its role in morphogenesis, contributes to virulence traits, including phagosome maturation blockade, host cell damages, and proangiogenic growth factor induction during interactions with hosts. IMPORTANCE Mucor is intrinsically resistant to most known antifungals, which makes mucormycosis treatment challenging. Calcineurin is a serine/threonine phosphatase that is widely conserved across eukaryotes. When calcineurin function is inhibited in Mucor, growth shifts to a less virulent yeast growth form, which makes calcineurin an attractive target for development of new antifungal drugs. Previously, we identified two distinct mechanisms through which Mucor can become resistant to calcineurin inhibitors involving Mendelian mutations in the gene for FKBP12, including mechanisms corresponding to calcineurin A or B subunits and epimutations silencing the FKBP12 gene. Here, we identified a third novel mechanism where loss-of-function mutations in the amino acid permease corresponding to the bycA gene contribute to resistance against calcineurin inhibitors. When calcineurin activity is absent, BycA can activate protein kinase A (PKA) to promote yeast growth via a cAMP-independent pathway. Our data also show that calcineurin activity contributes to host-pathogen interactions primarily in the pathogenesis of Mucor.
format	article
author	Sandeep Vellanki R. Blake Billmyre Alejandra Lorenzen Micaela Campbell Broderick Turner Eun Young Huh Joseph Heitman Soo Chan Lee
author_facet	Sandeep Vellanki R. Blake Billmyre Alejandra Lorenzen Micaela Campbell Broderick Turner Eun Young Huh Joseph Heitman Soo Chan Lee
author_sort	Sandeep Vellanki
title	A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
title_short	A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
title_full	A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
title_fullStr	A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
title_full_unstemmed	A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>
title_sort	novel resistance pathway for calcineurin inhibitors in the human-pathogenic mucorales <named-content content-type="genus-species">mucor circinelloides</named-content>
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/12849759825f4cdd96318e89c59485e8
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A Novel Resistance Pathway for Calcineurin Inhibitors in the Human-Pathogenic Mucorales <named-content content-type="genus-species">Mucor circinelloides</named-content>

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