scholarly journals The Aspergillus fumigatus P-Type Golgi Apparatus Ca2+/Mn2+ ATPase PmrA Is Involved in Cation Homeostasis and Cell Wall Integrity but Is Not Essential for Pathogenesis

2010 ◽  
Vol 9 (3) ◽  
pp. 472-476 ◽  
Author(s):  
Nadthanan Pinchai ◽  
Praveen Rao Juvvadi ◽  
Jarrod R. Fortwendel ◽  
B. Zachary Perfect ◽  
Luise E. Rogg ◽  
...  
Keyword(s):  
Cell Wall ◽  
Golgi Apparatus ◽  
Aspergillus Fumigatus ◽  
Cell Wall Integrity ◽  
Content Type ◽  
Growth Defects ◽  
Chitin Content ◽  
Cation Homeostasis ◽  
P Type ◽  
Calcineurin Pathway

ABSTRACT The Aspergillus fumigatus ΔpmrA (Golgi apparatus Ca2+/Mn2+ P-type ATPase) strain has osmotically suppressible basal growth defects and cationic tolerance associated with increased expression of calcineurin pathway genes. Despite increased β-glucan and chitin content, it is hypersensitive to cell wall inhibitors but remains virulent, suggesting a role for PmrA in cation homeostasis and cell wall integrity.

scholarly journals Putative PmrA and PmcA are important for normal growth, morphogenesis and cell wall integrity, but not for viability in Aspergillus nidulans

Microbiology ◽  
2014 ◽  
Vol 160 (11) ◽  
pp. 2387-2395 ◽  
Author(s):  
Hechun Jiang ◽  
Feifei Liu ◽  
Shizhu Zhang ◽  
Ling Lu
Keyword(s):  
Cell Wall ◽  
Plasma Membrane ◽  
Aspergillus Nidulans ◽  
Normal Growth ◽  
Cell Wall Integrity ◽  
Plasma Membrane Atpase ◽  
Growth Defects ◽  
Double Deletion ◽  
Intracellular Ca ◽  
P Type

P-type Ca2+-transporting ATPases are Ca2+ pumps, extruding cytosolic Ca2+ to the extracellular environment or the intracellular Ca2+ store lumens. In budding yeast, Pmr1 (plasma membrane ATPase related), and Pmc1 (plasma membrane calcium-ATPase) cannot be deleted simultaneously for it to survive in standard medium. Here, we deleted two putative Ca2+ pumps, designated AnPmrA and AnPmcA, from Aspergillus nidulans, and obtained the mutants ΔanpmrA and ΔanpmcA, respectively. Then, using ΔanpmrA as the starting strain, the promoter of its anpmcA was replaced with the alcA promoter to secure the mutant ΔanpmrAalcApmcA or its anpmcA was deleted completely to produce the mutant ΔanpmrAΔpmcA. Different from the case in Saccharomyces cerevisiae, double deletion of anpmrA and anpmcA was not lethal in A. nidulans. In addition, deletion of anpmrA and/or anpmcA had produced growth defects, although overexpression of AnPmc1 in ΔanpmrAalcApmcA could not restore the growth defects that resulted from the loss of AnPmrA. Moreover, we found AnPmrA was indispensable for maintenance of normal morphogenesis, especially in low-Ca2+/Mn2+ environments. Thus, our findings suggest AnPmrA and AnPmcA might play important roles in growth, morphogenesis and cell wall integrity in A. nidulans in a different way from that in yeasts.

scholarly journals Role of the Guanine Nucleotide Exchange Factor Rom2 in Cell Wall Integrity Maintenance of Aspergillus fumigatus

2012 ◽  
Vol 12 (2) ◽  
pp. 288-298 ◽  
Author(s):  
Sweta Samantaray ◽  
Michael Neubauer ◽  
Christoph Helmschrott ◽  
Johannes Wagener
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Guanine Nucleotide Exchange Factor ◽  
Cell Wall Integrity ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Content Type ◽  
Guanine Nucleotide Exchange ◽  
Stress Sensors ◽  
Nucleotide Exchange Factor

ABSTRACTAspergillus fumigatusis a mold and the causal agent of invasive aspergillosis, a systemic disease with high lethality. Recently, we identified and functionally characterized three stress sensors implicated in the cell wall integrity (CWI) signaling of this pathogen, namely, Wsc1, Wsc3, and MidA. Here, we functionally characterize Rom2, a guanine nucleotide exchange factor with essential function for the cell wall integrity ofA. fumigatus. A conditionalrom2mutant has severe growth defects under repressive conditions and incorporates all phenotypes of the three cell wall integrity sensor mutants, e.g., the echinocandin sensitivity of the Δwsc1mutant and the Congo red, calcofluor white, and heat sensitivity of the ΔmidAmutant. Rom2 interacts with Rho1 and shows a similar intracellular distribution focused at the hyphal tips. Our results place Rom2 between the cell surface stress sensors Wsc1, Wsc3, MidA, and Rho1 and their downstream effector mitogen-activated protein (MAP) kinase module Bck1-Mkk2-MpkA.

scholarly journals Caspofungin Treatment of Aspergillus fumigatus Results in ChsG-Dependent Upregulation of Chitin Synthesis and the Formation of Chitin-Rich Microcolonies

2015 ◽  
Vol 59 (10) ◽  
pp. 5932-5941 ◽  
Author(s):  
Louise A. Walker ◽  
Keunsook K. Lee ◽  
Carol A. Munro ◽  
Neil A. R. Gow
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Dynamic Imaging ◽  
Chitin Synthase ◽  
Response To Treatment ◽  
Chitin Synthesis ◽  
Calcofluor White ◽  
Content Type ◽  
Chitin Content

ABSTRACTTreatment ofAspergillus fumigatuswith echinocandins such as caspofungin inhibits the synthesis of cell wall β-1,3-glucan, which triggers a compensatory stimulation of chitin synthesis. Activation of chitin synthesis can occur in response to sub-MICs of caspofungin and to CaCl2and calcofluor white (CFW), agonists of the protein kinase C (PKC), and Ca2+-calcineurin signaling pathways.A. fumigatusmutants with thechsgene (encoding chitin synthase) deleted (ΔAfchs) were tested for their response to these agonists to determine the chitin synthase enzymes that were required for the compensatory upregulation of chitin synthesis. Only the ΔAfchsGmutant was hypersensitive to caspofungin, and all other ΔAfchsmutants tested remained capable of increasing their chitin content in response to treatment with CaCl2and CFW and caspofungin. The resulting increase in cell wall chitin content correlated with reduced susceptibility to caspofungin in the wild type and all ΔAfchsmutants tested, with the exception of the ΔAfchsGmutant, which remained sensitive to caspofungin.In vitroexposure to the chitin synthase inhibitor, nikkomycin Z, along with caspofungin demonstrated synergistic efficacy that was againAfChsG dependent. Dynamic imaging using microfluidic perfusion chambers demonstrated that treatment with sub-MIC caspofungin resulted initially in hyphal tip lysis. However, thickened hyphae emerged that formed aberrant microcolonies in the continued presence of caspofungin. In addition, intrahyphal hyphae were formed in response to echinocandin treatment. Thesein vitrodata demonstrate thatA. fumigatushas the potential to survive echinocandin treatmentin vivobyAfChsG-dependent upregulation of chitin synthesis. Chitin-rich cells may, therefore, persist in human tissues and act as the focus for breakthrough infections.

scholarly journals Heat Shock Protein 90 Is Required for Conidiation and Cell Wall Integrity in Aspergillus fumigatus

2012 ◽  
Vol 11 (11) ◽  
pp. 1324-1332 ◽  
Author(s):  
Frédéric Lamoth ◽  
Praveen R. Juvvadi ◽  
Jarrod R. Fortwendel ◽  
William J. Steinbach
Keyword(s):  
Cell Wall ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Aspergillus Fumigatus ◽  
Cellular Localization ◽  
Fluorescent Protein ◽  
Heat Shock Protein 90 ◽  
Cell Wall Integrity ◽  
Compensatory Mechanisms ◽  
Content Type

ABSTRACTHeat shock protein 90 (Hsp90) is a eukaryotic molecular chaperone. Its involvement in the resistance ofCandida albicansto azole and echinocandin antifungals is well established. However, little is known about Hsp90's function in the filamentous fungal pathogenAspergillus fumigatus. We investigated the role of Hsp90 inA. fumigatusby genetic repression and examined its cellular localization under various stress conditions. Failure to generate a deletion strain ofhsp90suggested that it is essential. Genetic repression of Hsp90 was achieved by an inducible nitrogen-dependent promoter (pniiA-Hsp90) and led to decreased spore viability, decreased hyphal growth, and severe defects in germination and conidiation concomitant with the downregulation of the conidiation-specific transcription factorsbrlA,wetA, andabaA. Hsp90 repression potentiated the effect of cell wall inhibitors affecting the β-glucan structure of the cell wall (caspofungin, Congo red) and of the calcineurin inhibitor FK506, supporting a role in regulating cell wall integrity pathways. Moreover, compromising Hsp90 abolished the paradoxical effect of caspofungin. Pharmacological inhibition of Hsp90 by geldanamycin and its derivatives (17-AAG and 17-DMAG) resulted in similar effects. C-terminal green fluorescent protein (GFP) tagging of Hsp90 revealed mainly cytosolic distribution under standard growth conditions. However, treatment with caspofungin resulted in Hsp90 accumulation at the cell wall and at sites of septum formation, further highlighting its role in cell wall stress compensatory mechanisms. Targeting Hsp90 with fungal-specific inhibitors to cripple stress response compensatory pathways represents an attractive new antifungal strategy.

scholarly journals The Aspergillus fumigatus sitA Phosphatase Homologue Is Important for Adhesion, Cell Wall Integrity, Biofilm Formation, and Virulence

2015 ◽  
Vol 14 (8) ◽  
pp. 728-744 ◽  
Author(s):  
Vinícius Leite Pedro Bom ◽  
Patrícia Alves de Castro ◽  
Lizziane K. Winkelströter ◽  
Marçal Marine ◽  
Juliana I. Hori ◽  
...  
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Biofilm Formation ◽  
Essential Gene ◽  
Pathogenic Fungus ◽  
Invasive Pulmonary Aspergillosis ◽  
Cell Wall Integrity ◽  
Necrosis Factor Alpha ◽  
Content Type ◽  
Opportunistic Pathogenic

ABSTRACTAspergillus fumigatusis an opportunistic pathogenic fungus able to infect immunocompromised patients, eventually causing disseminated infections that are difficult to control and lead to high mortality rates. It is important to understand how the signaling pathways that regulate these factors involved in virulence are orchestrated. Protein phosphatases are central to numerous signal transduction pathways. Here, we characterize theA. fumigatusprotein phosphatase 2A SitA, theSaccharomyces cerevisiaeSit4p homologue. ThesitAgene is not an essential gene, and we were able to construct anA. fumigatusnull mutant. The ΔsitAstrain had decreased MpkA phosphorylation levels, was more sensitive to cell wall-damaging agents, had increased β-(1,3)-glucan and chitin, was impaired in biofilm formation, and had decreased protein kinase C activity. The ΔsitAstrain is more sensitive to several metals and ions, such as MnCl2, CaCl2, and LiCl, but it is more resistant to ZnSO4. The ΔsitAstrain was avirulent in a murine model of invasive pulmonary aspergillosis and induces an augmented tumor necrosis factor alpha (TNF-α) response in mouse macrophages. These results stress the importance ofA. fumigatusSitA as a possible modulator of PkcA/MpkA activity and its involvement in the cell wall integrity pathway.

scholarly journals Putative Membrane Receptors Contribute to Activation and Efficient Signaling of Mitogen-Activated Protein Kinase Cascades during Adaptation of Aspergillus fumigatus to Different Stressors and Carbon Sources

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Lilian Pereira Silva ◽  
Dean Frawley ◽  
Leandro José de Assis ◽  
Ciara Tierney ◽  
Alastair B. Fleming ◽  
...  
Keyword(s):  
Cell Wall ◽  
Protein Kinase ◽  
Aspergillus Fumigatus ◽  
Fungal Pathogen ◽  
Cell Wall Integrity ◽  
Mitogen Activated Protein Kinase ◽  
Hog Pathway ◽  
Content Type ◽  
High Osmolarity Glycerol ◽  
Mitogen Activated Protein

ABSTRACT The high-osmolarity glycerol (HOG) response pathway is a multifunctional signal transduction pathway that specifically transmits ambient osmotic signals. Saccharomyces cerevisiae Hog1p has two upstream signaling branches, the sensor histidine kinase Sln1p and the receptor Sho1p. The Sho1p branch includes two other proteins, the Msb2p mucin and Opy2p. Aspergillus fumigatus is the leading cause of pulmonary fungal diseases. Here, we investigated the roles played by A. fumigatus SlnASln1p, ShoASho1p, MsbAMsb2p, and OpyAOpy2p putative homologues during the activation of the mitogen-activated protein kinase (MAPK) HOG pathway. The shoA, msbA, and opyA singly and doubly null mutants are important for the cell wall integrity (CWI) pathway, oxidative stress, and virulence as assessed by a Galleria mellonella model. Genetic interactions of ShoA, MsbA, and OpyA are also important for proper activation of the SakAHog1p and MpkASlt2 cascade and the response to osmotic and cell wall stresses. Comparative label-free quantitative proteomics analysis of the singly null mutants with the wild-type strain upon caspofungin exposure indicates that the absence of ShoA, MsbA, and OpyA affects the osmotic stress response, carbohydrate metabolism, and protein degradation. The putative receptor mutants showed altered trehalose and glycogen accumulation, suggesting a role for ShoA, MsbA, and OpyA in sugar storage. Protein kinase A activity was also decreased in these mutants. We also observed genetic interactions between SlnA, ShoA, MsbA, and OpyA, suggesting that both branches are important for activation of the HOG/CWI pathways. Our results help in the understanding of the activation and modulation of the HOG and CWI pathways in this important fungal pathogen. IMPORTANCE Aspergillus fumigatus is an important human-pathogenic fungal species that is responsible for a high incidence of infections in immunocompromised individuals. A. fumigatus high-osmolarity glycerol (HOG) and cell wall integrity pathways are important for the adaptation to different forms of environmental adversity such as osmotic and oxidative stresses, nutrient limitations, high temperatures, and other chemical and mechanical stresses that may be produced by the host immune system and antifungal drugs. Little is known about how these pathways are activated in this fungal pathogen. Here, we characterize four A. fumigatus putative homologues that are important for the activation of the yeast HOG pathway. A. fumigatus SlnASln1p, ShoASho1p, MsbAMsb2p, and OpyAOpy2p are genetically interacting and are essential for the activation of the HOG and cell wall integrity pathways. Our results contribute to the understanding of A. fumigatus adaptation to the host environment.

scholarly journals The Cell Wall Integrity Pathway Contributes to the Early Stages of Aspergillus fumigatus Asexual Development

2020 ◽  
Vol 86 (7) ◽  
Author(s):  
Marina Campos Rocha ◽  
João Henrique Tadini Marilhano Fabri ◽  
Isabelle Taira Simões ◽  
Rafael Silva-Rocha ◽  
Daisuke Hagiwara ◽  
...  
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Filamentous Fungi ◽  
Cell Wall Integrity ◽  
Asexual Development ◽  
Remarkable Feature ◽  
Susceptible Host ◽  
Content Type ◽  
Cell Wall Integrity Pathway ◽  
Cell Wall Remodeling

ABSTRACT Aspergillus fumigatus is a major cause of human disease. The survival of this fungus is dependent on the cell wall organization and function of its components. The cell wall integrity pathway (CWIP) is the primary signaling cascade that controls de novo synthesis of the cell wall in fungi. Abundant conidiation is a hallmark in A. fumigatus, and uptake of conidia by a susceptible host is usually the initial event in infection. The formation of conidia is mediated by the development of fungus-specific specialized structures, conidiophores, which are accompanied by cell wall remodeling. The molecular regulation of these changes in cell wall composition required for the rise of conidiophore from the solid surface and to disperse the conidia into the air is currently unknown. Here, we investigated the role of CWIP in conidiation. We show that CWIP pkcAG579R, ΔmpkA, and ΔrlmA mutants displayed reduced conidiation during synchronized asexual differentiation. The transcription factor RlmA directly regulated the expression of regulators of conidiation, including flbB, flbC, brlA, abaA, and rasB, as well as genes involved in cell wall synthesis and remodeling, and this affected the chitin content in aerial hyphae. Phosphorylation of RlmA and MpkA was increased during asexual differentiation. We also observed that MpkA physically associated with the proteins FlbB, FlbC, BrlA, and RasB during this process, suggesting another level of cross talk between the CWIP and asexual development pathways. In summary, our results support the conclusion that one function of the CWIP is the regulation of asexual development in filamentous fungi. IMPORTANCE A remarkable feature of the human pathogen Aspergillus fumigatus is its ability to produce impressive amounts of infectious propagules known as conidia. These particles reach immunocompromised patients and may initiate a life-threatening mycosis. The conidiation process in Aspergillus is governed by a sequence of proteins that coordinate the development of conidiophores. This process requires the remodeling of the cell wall so that the conidiophores can rise and withstand the chains of conidia. The events regulating cell wall remodeling during conidiation are currently unknown. Here, we show that the cell wall integrity pathway (CWIP) components RlmA and MpkA directly contribute to the activation of the conidiation cascade by enabling transcription or phosphorylation of critical proteins involved in asexual development. This study points to an essential role for the CWIP during conidiation and provides further insights into the complex regulation of asexual development in filamentous fungi.

scholarly journals The Aspergillus fumigatus Mucin MsbA Regulates the Cell Wall Integrity Pathway and Controls Recognition of the Fungus by the Immune System

mSphere ◽  
2019 ◽  
Vol 4 (3) ◽  
Author(s):  
Isabella Luísa da Silva Gurgel ◽  
Karina Talita de Oliveira Santana Jorge ◽  
Nathália Luísa Sousa de Oliveira Malacco ◽  
Jéssica Amanda Marques Souza ◽  
Marina Campos Rocha ◽  
...  
Keyword(s):  
Cell Wall ◽  
Immune System ◽  
Aspergillus Fumigatus ◽  
Fungal Growth ◽  
Cell Wall Composition ◽  
Cell Wall Integrity ◽  
Cell Physiology ◽  
Content Type ◽  
Cell Wall Morphogenesis

ABSTRACT Aspergillus fumigatus is a filamentous fungus which causes invasive pulmonary aspergillosis in immunocompromised individuals. In fungi, cell signaling and cell wall plasticity are crucial for maintaining physiologic processes. In this context, Msb2 is an important signaling mucin responsible for activation of a variety of mitogen-activated protein kinase (MAPK)-dependent signaling pathways that regulate cell growth in several organisms, such as the cell wall integrity (CWI) pathway. Here, we aimed to characterize the MSB2 homologue in A. fumigatus. Our results showed that MsbA plays a role in the vegetative and reproductive development of the fungus, in stress adaptation, and in resistance to antifungal drugs by modulating the CWI pathway gene expression. Importantly, cell wall composition is also responsible for activation of diverse receptors of the host immune system, thus leading to a proper immune response. In a model of acute Aspergillus pulmonary infection, results demonstrate that the ΔmsbA mutant strain induced less inflammation with diminished cell influx into the lungs and lower cytokine production, culminating in increased lethality rate. These results characterize for the first time the role of the signaling mucin MsbA in the pathogen A. fumigatus, as a core sensor for cell wall morphogenesis and an important regulator of virulence. IMPORTANCE Aspergillus fumigatus is an opportunistic fungus with great medical importance. During infection, Aspergillus grows, forming hyphae that colonize the lung tissue and invade and spread over the mammal host, resulting in high mortality rates. The knowledge of the mechanisms responsible for regulation of fungal growth and virulence comprises an important point to better understand fungal physiology and host-pathogen interactions. Msb2 is a mucin that acts as a sensor and an upstream regulator of the MAPK pathway responsible for fungal development in Candida albicans and Aspergillus nidulans. Here, we show the role of the signaling mucin MsbA in the pathogen A. fumigatus, as a core sensor for cell wall morphogenesis, fungal growth, and virulence. Moreover, we show that cell wall composition, controlled by MsbA, is detrimental for fungal recognition and clearance by immune cells. Our findings are important for the understanding of how fungal sensors modulate cell physiology.

scholarly journals α1,3 Glucans Are Dispensable in Aspergillus fumigatus

2011 ◽  
Vol 11 (1) ◽  
pp. 26-29 ◽  
Author(s):  
Christine Henry ◽  
Jean-Paul Latgé ◽  
Anne Beauvais
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Vegetative Growth ◽  
Conidial Germination ◽  
Glucan Synthase ◽  
Content Type ◽  
Chitin Content ◽  
A Cell

ABSTRACTA triple α1,3 glucan synthase mutant ofAspergillus fumigatusobtained by successive deletions of the three α1,3 glucan synthase genes (AGS1,AGS2, andAGS3) has a cell wall devoid of α1,3 glucans. The lack of α1,3 glucans affects neither conidial germination nor mycelial vegetative growth and is compensated by an increase in β1,3 glucan and/or chitin content.

scholarly journals Differential Regulation of the Cell Wall Integrity Mitogen-Activated Protein Kinase Pathway in Budding Yeast by the Protein Tyrosine Phosphatases Ptp2 and Ptp3

1999 ◽  
Vol 19 (11) ◽  
pp. 7651-7660 ◽  
Author(s):  
Christopher P. Mattison ◽  
Scott S. Spencer ◽  
Kurt A. Kresge ◽  
Ji Lee ◽  
Irene M. Ota
Keyword(s):  
Cell Wall ◽  
Protein Tyrosine Phosphatases ◽  
Negative Regulator ◽  
Cell Wall Integrity ◽  
Dependent Manner ◽  
Tyrosine Phosphatases ◽  
Protein Tyrosine ◽  
Growth Defects ◽  
Mitogen Activated Protein

ABSTRACT Mitogen-activated protein kinases (MAPKs) are inactivated by dual-specificity and protein tyrosine phosphatases (PTPs) in yeasts. InSaccharomyces cerevisiae, two PTPs, Ptp2 and Ptp3, inactivate the MAPKs, Hog1 and Fus3, with different specificities. To further examine the functions and substrate specificities of Ptp2 and Ptp3, we tested whether they could inactivate a third MAPK, Mpk1, in the cell wall integrity pathway. In vivo and in vitro evidence indicates that both PTPs inactivate Mpk1, but Ptp2 is the more effective negative regulator. Multicopy expression of PTP2, but not PTP3, suppressed growth defects due to the MEK kinase mutation, BCK1-20, and the MEK mutation,MKK1-386, that hyperactivate this pathway. In addition, deletion of PTP2, but not PTP3, exacerbated growth defects due to MKK1-386. Other evidence supported a role for Ptp3 in this pathway. Expression of MKK1-386 was lethal in the ptp2Δ ptp3Δ strain but not in either single PTP deletion strain. In addition, the ptp2Δ ptp3Δ strain showed higher levels of heat stress-induced Mpk1-phosphotyrosine than the wild-type strain or strains lacking either PTP. The PTPs also showed differences in vitro. Ptp2 was more efficient than Ptp3 at binding and dephosphorylating Mpk1. Another factor that may contribute to the greater effectiveness of Ptp2 is its subcellular localization. Ptp2 is predominantly nuclear whereas Ptp3 is cytoplasmic, suggesting that active Mpk1 is present in the nucleus. Last, PTP2 but not PTP3 transcript increased in response to heat shock in a Mpk1-dependent manner, suggesting that Ptp2 acts in a negative feedback loop to inactivate Mpk1.

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