scholarly journals Identification and Phenotypic Characterization of Hsp90 Phosphorylation Sites That Modulate Virulence Traits in the Major Human Fungal Pathogen Candida albicans

2021 ◽  
Vol 11 ◽  
Author(s):  
Leenah Alaalm ◽  
Julia L. Crunden ◽  
Mark Butcher ◽  
Ulrike Obst ◽  
Ryann Whealy ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Environmental Stress ◽  
Stress Responses ◽  
Fungal Pathogens ◽  
Phosphorylation Site ◽  
Pathogenic Fungi ◽  
Phenotypic Characterization ◽  
Fungal Virulence ◽  
Post Translational Modifications ◽  
Virulence Traits

The highly conserved, ubiquitous molecular chaperone Hsp90 is a key regulator of cellular proteostasis and environmental stress responses. In human pathogenic fungi, which kill more than 1.6 million patients each year worldwide, Hsp90 governs cellular morphogenesis, drug resistance, and virulence. Yet, our understanding of the regulatory mechanisms governing fungal Hsp90 function remains sparse. Post-translational modifications are powerful components of nature’s toolbox to regulate protein abundance and function. Phosphorylation in particular is critical in many cellular signaling pathways and errant phosphorylation can have dire consequences for the cell. In the case of Hsp90, phosphorylation affects its stability and governs its interactions with co-chaperones and clients. Thereby modulating the cell’s ability to cope with environmental stress. Candida albicans, one of the leading human fungal pathogens, causes ~750,000 life-threatening invasive infections worldwide with unacceptably high mortality rates. Yet, it remains unknown if and how Hsp90 phosphorylation affects C. albicans virulence traits. Here, we show that phosphorylation of Hsp90 is critical for expression of virulence traits. We combined proteomics, molecular evolution analyses and structural modeling with molecular biology to characterize the role of Hsp90 phosphorylation in this non-model pathogen. We demonstrated that phosphorylation negatively affects key virulence traits, such as the thermal stress response, morphogenesis, and drug susceptibility. Our results provide the first record of a specific Hsp90 phosphorylation site acting as modulator of fungal virulence. Post-translational modifications of Hsp90 could prove valuable in future exploitations as antifungal drug targets.

scholarly journals Transcriptome analysis uncovers a link between copper metabolism, and both fungal fitness and antifungal sensitivity in the opportunistic yeast Candida albicans

2020 ◽  
Author(s):  
Inès Khemiri ◽  
Faiza Tebbji ◽  
Adnane Sellam
Keyword(s):  
Candida Albicans ◽  
Stress Responses ◽  
Copper Metabolism ◽  
Pathogenic Fungi ◽  
Global Gene Expression ◽  
Host Cells ◽  
Fungal Virulence ◽  
Copper Transporter ◽  
Gene Expression Dynamic ◽  
As Stress

AbstractCopper homeostasis is an important determinant for virulence of many human pathogenic fungi such as the highly prevalent yeast Candida albicans. However, beyond the copper transporter Ctr1, little is known regarding other genes and biological processes that are affected by copper. To gain insight into the cellular processes that are modulated by copper abundance in C. albicans, we monitored the global gene expression dynamic under both copper depletion and excess using RNA-seq. Beyond copper metabolism, other different transcriptional programs related to fungal fitness such as stress responses, antifungal sensitivity, host invasion and commensalism were modulated in response to copper variations. We have also investigated the transcriptome of the mutant of the copper utilization regulator, mac1, and identified potential direct targets of this transcription factor under copper starvation. We also showed that Mac1 was required for the invasion and adhesion to host cells and antifungal tolerance. This study provides a framework for future studies to examine the link between copper metabolism and essential functions that modulate fungal virulence and fitness inside the host.

scholarly journals A novel Hsp90 phospho-switch modulates virulence in the major human fungal pathogen Candida albicans

2020 ◽  
Author(s):  
Leenah Alaalm ◽  
Julia L. Crunden ◽  
Mark Butcher ◽  
Ulrike Obst ◽  
Ryann Whealy ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Stress Responses ◽  
Fungal Pathogen ◽  
Drug Targets ◽  
Fungal Infections ◽  
Pathogenic Fungi ◽  
First Record ◽  
Drug Susceptibility ◽  
Phosphorylation Sites ◽  
Fungal Virulence

The ubiquitous molecular chaperone Hsp90 is a key regulator of cellular proteostasis and environmental stress responses. Hsp90 also regulates cellular morphogenesis, drug resistance, and virulence in human pathogenic fungi, which kill more than 1.6 million patients each year worldwide. Invasive fungal infections are difficult to treat due to the lack of effective antifungal therapies, resulting in mortality rates of up to 95%. As a key regulator of fungal virulence, Hsp90 is an attractive therapeutic target. However, fungal and animal homologs are highly conserved, impeding fungal-specific targeting. Thus, understanding the factors that regulate Hsp90 could provide an alternative strategy aimed at exclusively targeting this regulator of fungal virulence. Here, we demonstrate how CK2-mediated phosphorylation of two Hsp90 residues modulates virulence in a major fungal pathogen of humans, Candida albicans. We combined proteomics, molecular evolution and structural modelling with molecular biology to identify and characterize two Hsp90 phosphorylation sites. Phosphorylation negatively affects thermal stress response, morphogenesis, drug susceptibility and fungal virulence. Our results provide the first record of specific Hsp90 phosphorylation sites acting as modulators of fungal virulence. Post-translational modifications of Hsp90 could prove valuable in future exploitation as antifungal drug targets.

scholarly journals The Ustilago hordei–Barley Interaction is a Versatile System for Characterization of Fungal Effectors

2021 ◽  
Vol 7 (2) ◽  
pp. 86
Author(s):  
Bilal Ökmen ◽  
Daniela Schwammbach ◽  
Guus Bakkeren ◽  
Ulla Neumann ◽  
Gunther Doehlemann
Keyword(s):  
Fungal Pathogens ◽  
Plant Pathogens ◽  
Expression System ◽  
Pathogenic Fungi ◽  
Effector Proteins ◽  
Mating Partner ◽  
Fungal Virulence ◽  
Functional Studies ◽  
Infection Structures ◽  
Ustilago Hordei

Obligate biotrophic fungal pathogens, such as Blumeria graminis and Puccinia graminis, are amongst the most devastating plant pathogens, causing dramatic yield losses in many economically important crops worldwide. However, a lack of reliable tools for the efficient genetic transformation has hampered studies into the molecular basis of their virulence or pathogenicity. In this study, we present the Ustilago hordei–barley pathosystem as a model to characterize effectors from different plant pathogenic fungi. We generate U. hordei solopathogenic strains, which form infectious filaments without the presence of a compatible mating partner. Solopathogenic strains are suitable for heterologous expression system for fungal virulence factors. A highly efficient Crispr/Cas9 gene editing system is made available for U. hordei. In addition, U. hordei infection structures during barley colonization are analyzed using transmission electron microscopy, showing that U. hordei forms intracellular infection structures sharing high similarity to haustoria formed by obligate rust and powdery mildew fungi. Thus, U. hordei has high potential as a fungal expression platform for functional studies of heterologous effector proteins in barley.

scholarly journals Novel Antifungal Compounds Discovered in Medicines for Malaria Venture’s Malaria Box

mSphere ◽  
2018 ◽  
Vol 3 (2) ◽  
Author(s):  
Eric H. Jung ◽  
David J. Meyers ◽  
Jürgen Bosch ◽  
Arturo Casadevall
Keyword(s):  
Candida Albicans ◽  
Antifungal Activity ◽  
Cryptococcus Neoformans ◽  
Fungal Pathogens ◽  
Pathogenic Fungi ◽  
Cryptococcus Gattii ◽  
Antifungal Drugs ◽  
Resistant Bacteria ◽  
Animal Cells ◽  
Malaria Box

ABSTRACTSimilarities in fungal and animal cells make antifungal discovery efforts more difficult than those for other classes of antimicrobial drugs. Currently, there are only three major classes of antifungal drugs used for the treatment of systemic fungal diseases: polyenes, azoles, and echinocandins. Even in situations where the offending fungal organism is susceptible to the available drugs, treatment courses can be lengthy and unsatisfactory, since eradication of infection is often very difficult, especially in individuals with impaired immunity. Consequently, there is a need for new and more effective antifungal drugs. We have identified compounds with significant antifungal activity in the Malaria Box (Medicines for Malaria Ventures, Geneva, Switzerland) that have higher efficacy than some of the currently used antifungal drugs. Our best candidate, MMV665943 (IUPAC name 4-[6-[[2-(4-aminophenyl)-3H-benzimidazol-5-yl]methyl]-1H-benzimidazol-2-yl]aniline), here referred to as DM262, showed 16- to 32-fold-higher activity than fluconazole againstCryptococcus neoformans. There was also significant antifungal activity in other fungal species with known antifungal resistance, such asLomentospora prolificansandCryptococcus gattii. Antifungal activity was also observed against a common fungus,Candida albicans. These results are important because they offer a potentially new class of antifungal drugs and the repurposing of currently available therapeutics.IMPORTANCEMuch like the recent increase in drug-resistant bacteria, there is a rise in antifungal-resistant strains of pathogenic fungi. There is a need for novel and more potent antifungal therapeutics. Consequently, we investigated a mixed library of drug-like and probe-like compounds with activity inPlasmodiumspp. for activity against two common fungal pathogens,Cryptococcus neoformansandCandida albicans, along with two less common pathogenic species,Lomentospora prolificansandCryptococcus gattii. We uncover a previously uncharacterized drug with higher broad-spectrum antifungal activity than some current treatments. Our findings may eventually lead to a compound added to the arsenal of antifungal therapeutics.

scholarly journals Structure-guided approaches to targeting stress responses in human fungal pathogens

2020 ◽  
Vol 295 (42) ◽  
pp. 14458-14472
Author(s):  
Emmanuelle V. LeBlanc ◽  
Elizabeth J. Polvi ◽  
Amanda O. Veri ◽  
Gilbert G. Privé ◽  
Leah E. Cowen
Keyword(s):  
Stress Response ◽  
Stress Responses ◽  
Fungal Pathogens ◽  
Fungal Infections ◽  
Fungal Growth ◽  
Small Gtpase ◽  
Ecological Niches ◽  
Attractive Alternative ◽  
Response Regulators ◽  
Fungal Virulence

Fungi inhabit extraordinarily diverse ecological niches, including the human body. Invasive fungal infections have a devastating impact on human health worldwide, killing ∼1.5 million individuals annually. The majority of these deaths are attributable to species of Candida, Cryptococcus, and Aspergillus. Treating fungal infections is challenging, in part due to the emergence of resistance to our limited arsenal of antifungal agents, necessitating the development of novel therapeutic options. Whereas conventional antifungal strategies target proteins or cellular components essential for fungal growth, an attractive alternative strategy involves targeting proteins that regulate fungal virulence or antifungal drug resistance, such as regulators of fungal stress responses. Stress response networks enable fungi to adapt, grow, and cause disease in humans and include regulators that are highly conserved across eukaryotes as well as those that are fungal-specific. This review highlights recent developments in elucidating crystal structures of fungal stress response regulators and emphasizes how this knowledge can guide the design of fungal-selective inhibitors. We focus on the progress that has been made with highly conserved regulators, including the molecular chaperone Hsp90, the protein phosphatase calcineurin, and the small GTPase Ras1, as well as with divergent stress response regulators, including the cell wall kinase Yck2 and trehalose synthases. Exploring structures of these important fungal stress regulators will accelerate the design of selective antifungals that can be deployed to combat life-threatening fungal diseases.

scholarly journals Comparative Genomics and the Evolution of Pathogenicity in Human Pathogenic Fungi

2010 ◽  
Vol 10 (1) ◽  
pp. 34-42 ◽  
Author(s):  
Gary P. Moran ◽  
David C. Coleman ◽  
Derek J. Sullivan
Keyword(s):  
Fungal Pathogens ◽  
Direct Interaction ◽  
Pathogenic Fungi ◽  
Gene Families ◽  
Fungal Pathogenesis ◽  
Individual Species ◽  
Fungal Virulence ◽  
Content Type ◽  
Sequencing Technologies ◽  
Genes Encoding

ABSTRACTBecause most fungi have evolved to be free-living in the environment and because the infections they cause are usually opportunistic in nature, it is often difficult to identify specific traits that contribute to fungal pathogenesis. In recent years, there has been a surge in the number of sequenced genomes of human fungal pathogens, and comparison of these sequences has proved to be an excellent resource for exploring commonalities and differences in how these species interact with their hosts. In order to survive in the human body, fungi must be able to adapt to new nutrient sources and environmental stresses. Therefore, genes involved in carbohydrate and amino acid metabolism and transport and genes encoding secondary metabolites tend to be overrepresented in pathogenic species (e.g.,Aspergillus fumigatus). However, it is clear that human commensal yeast species such asCandida albicanshave also evolved a range of specific factors that facilitate direct interaction with host tissues. The evolution of virulence across the human pathogenic fungi has occurred largely through very similar mechanisms. One of the most important mechanisms is gene duplication and the expansion of gene families, particularly in subtelomeric regions. Unlike the case for prokaryotic pathogens, horizontal transfer of genes between species and other genera does not seem to have played a significant role in the evolution of fungal virulence. New sequencing technologies promise the prospect of even greater numbers of genome sequences, facilitating the sequencing of multiple genomes and transcriptomes within individual species, and will undoubtedly contribute to a deeper insight into fungal pathogenesis.

scholarly journals The Heat-Induced Molecular Disaggregase Hsp104 of Candida albicans Plays a Role in Biofilm Formation and Pathogenicity in a Worm Infection Model

2012 ◽  
Vol 11 (8) ◽  
pp. 1012-1020 ◽  
Author(s):  
Alessandro Fiori ◽  
Soňa Kucharíková ◽  
Gilmer Govaert ◽  
Bruno P. A. Cammue ◽  
Karin Thevissen ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Stress Responses ◽  
Biofilm Formation ◽  
Fungal Pathogens ◽  
Elevated Temperatures ◽  
Structural Defects ◽  
Infection Model ◽  
Content Type

ABSTRACT The consequences of deprivation of the molecular chaperone Hsp104 in the fungal pathogen Candida albicans were investigated. Mutants lacking HSP104 became hypersusceptible to lethally high temperatures, similarly to the corresponding mutants of Saccharomyces cerevisiae , whereas normal susceptibility was restored upon reintroduction of the gene. By use of a strain whose only copy of HSP104 is an ectopic gene under the control of a tetracycline-regulated promoter, expression of Hsp104 prior to the administration of heat shock could be demonstrated to be sufficient to confer protection from the subsequent temperature increase. This result points to a key role for Hsp104 in orchestrating the cell response to elevated temperatures. Despite their not showing evident growth or morphological defects, biofilm formation by cells lacking HSP104 proved to be defective in two established in vitro models that use polystyrene and polyurethane as the substrates. Biofilms formed by the wild-type and HSP104 -reconstituted strains showed patterns of intertwined hyphae in the extracellular matrix. In contrast, biofilm formed by the hsp104 Δ/ hsp104 Δ mutant showed structural defects and appeared patchy and loose. Decreased virulence of the hsp104 Δ/ hsp104 Δ mutant was observed in the Caenorhabditis elegans infection model, in which high in vivo temperature does not play a role. In agreement with the view that stress responses in fungal pathogens may have evolved to provide niche-specific adaptation to environmental conditions, these results provide an indication of a temperature-independent role for Hsp104 in support of Candida albicans virulence, in addition to its key role in governing thermotolerance.

scholarly journals Interplay between Candida albicans and the Antimicrobial Peptide Armory

2014 ◽  
Vol 13 (8) ◽  
pp. 950-957 ◽  
Author(s):  
Marc Swidergall ◽  
Joachim F. Ernst
Keyword(s):  
Candida Albicans ◽  
Defense Mechanisms ◽  
Fungal Pathogens ◽  
Pathogenic Fungi ◽  
Resistance Mechanisms ◽  
Immune Defense ◽  
Basal Resistance ◽  
Content Type ◽  
Normal Human ◽  
Molecular Patterns

ABSTRACTAntimicrobial peptides (AMPs) are key elements of innate immunity, which can directly kill multiple bacterial, viral, and fungal pathogens. The medically important fungusCandida albicanscolonizes different host niches as part of the normal human microbiota. Proliferation ofC. albicansis regulated through a complex balance of host immune defense mechanisms and fungal responses. Expression of AMPs against pathogenic fungi is differentially regulated and initiated by interactions of a variety of fungal pathogen-associated molecular patterns (PAMPs) with pattern recognition receptors (PRRs) on human cells. Inflammatory signaling and other environmental stimuli are also essential to control fungal proliferation and to prevent parasitism. To persist in the host,C. albicanshas developed a three-phase AMP evasion strategy, including secretion of peptide effectors, AMP efflux pumps, and regulation of signaling pathways. These mechanisms preventC. albicansfrom the antifungal activity of the major AMP classes, including cathelicidins, histatins, and defensins leading to a basal resistance. This minireview summarizes human AMP attack andC. albicansresistance mechanisms and current developments in the use of AMPs as antifungal agents.

scholarly journals Ploidy determines the consequences of antifungal-induced mutagenesis in Candida albicans, a human fungal pathogen

2019 ◽  
Author(s):  
Ognenka Avramovska ◽  
Meleah A. Hickman
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Environmental Stress ◽  
Genome Size ◽  
Fungal Pathogen ◽  
Fungal Pathogens ◽  
Antifungal Drugs ◽  
Relevant Model ◽  
Human Fungal Pathogen ◽  
Induced Mutagenesis

AbstractOrganismal ploidy state and environmental stress impact the mutational spectrum and the mutational rate. The human fungal pathogen Candida albicans, serves as a clinically relevant model for studying the interaction between eukaryotic ploidy and stress-induced mutagenesis. In this study, we compared the rates and types of genome perturbations in diploid and tetraploid C. albicans following exposure to two classes of antifungal drugs, azoles and echinocandins. We measured mutations at three different scales: point mutation, loss-of-heterozygosity (LOH), and genome size changes in cells treated with fluconazole and caspofungin. We find that caspofungin induced higher rates of mutation than fluconazole, likely an indirect result from the stress associated with cell wall perturbations rather than an inherent genotoxicity. Furthermore, we found disproportionately elevated rates of LOH and genome size changes in response to both antifungals in tetraploid C. albicans compared to diploid C. albicans, suggesting that the magnitude of stress-induced mutagenesis results from an interaction between ploidy state and the environment. These results have both clinical and evolutionary implications for how fungal pathogens generate mutations in response to antifungal drug stress, and may facilitate the emergence of antifungal resistance.

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