Aspergillus Fumigatus Cell Wall Research Articles

The basic leucine zipper transcription factor MeaB is critical for biofilm formation, cell wall integrity, and virulence in Aspergillus fumigatus.

The regulation of fungal cell wall biosynthesis is crucial for cell wall integrity maintenance and directly impacts fungal pathogen virulence. Although numerous genes are involved in fungal cell wall polysaccharide biosynthesis through multiple pathways, the underlying regulatory mechanism is still not fully understood. In this study, we identified and functionally characterized a direct downstream target of SomA, the basic-region leucine zipper transcription factor MeaB, playing a certain role in Aspergillus fumigatus cell wall integrity. Loss of meaB reduces hyphal growth, causes severe defects in galactosaminogalactan-mediated biofilm formation, and attenuates virulence in a Galleria mellonella infection model. Furthermore, the meaB null mutant strain exhibited hypersensitivity to cell wall-perturbing agents and significantly alters the cell wall structure. Transcriptional profile analysis revealed that MeaB positively regulates the expression of the galactosaminogalactan biosynthesis and β-1,3-glucanosyltransferase genes uge3, agd3, and sph3 and gel1, gel5, and gel7, respectively, as well as genes involved in amino sugar and nucleotide sugar metabolism. Further study demonstrated that MeaB could respond to cell wall stress and contribute to the proper expression of mitogen-activated protein kinase genes mpkA and mpkC in the presence of different concentrations of congo red. In conclusion, A. fumigatus MeaB plays a critical role in cell wall integrity by governing the expression of genes encoding cell wall-related proteins, thus impacting the virulence of this fungus.IMPORTANCEAspergillus fumigatus is a common opportunistic mold that causes life-threatening infections in immunosuppressed patients. The fungal cell wall is a complex and dynamic organelle essential for the development of pathogenic fungi. Genes involved in cell wall polysaccharide biosynthesis and remodeling are crucial for fungal pathogen virulence. However, the potential regulatory mechanism for cell wall integrity remains to be fully defined in A. fumigatus. In the present study, we identify basic-region leucine zipper transcription factor MeaB as an important regulator of cell wall galactosaminogalactan biosynthesis and β-1,3-glucan remodeling that consequently impacts stress response and virulence of fungal pathogens. Thus, we illuminate a mechanism of transcriptional control fungal cell wall polysaccharide biosynthesis and stress response. As these cell wall components are promising therapeutic targets for fungal infections, understanding the regulatory mechanism of such polysaccharides will provide new therapeutic opportunities.

Amino sugars influence Aspergillus fumigatus cell wall polysaccharide biosynthesis, and biofilm formation through interfering galactosaminogalactan deacetylation

Aspergillus fumigatus is a ubiquitous fungal pathogen responsible for a significant number of deaths annually due to invasive aspergillosis infection. While the utilization of diverse carbon sources, including amino sugars, has been explored in other fungi, its impact on A. fumigatus remains uncharted territory. In this study, we investigated A. fumigatus responses to glucose (Glc), glucosamine (GlcN) and N-acetylglucosamine (GlcNAc) as carbon sources. GlcN inhibited growth, reduced sporulation and delayed germination, while GlcNAc had no such effects. Both amino sugars induced alterations in cell wall composition, leading to a reduction in glucan and galactomannan levels while increasing chitin and mannan content, rendering A. fumigatus susceptible to cell wall stress and osmotic stress. GlcN repressed biofilm formation via downregulation of galactosaminogalactan (GAG) cluster genes, notably agd3, which encodes a GAG-specific deacetylase. Moreover, GlcN increased biofilm susceptibility to echinocandins, suggesting its potential for enhancing the effectiveness of antifungal treatments. This study sheds light on the multifaceted effects of amino sugars on A. fumigatus, encompassing growth, cell wall biosynthesis, and biofilm formation, offering promising avenues for innovative aspergillosis treatment strategies.

Cell wall of Aspergillus fumigatus: Variability and response to stress

The fungal cell is surrounded by a thick cell wall which obviously play an essential role in the protection of the fungus against external aggressive environments. In spite of 50 years of studies, the cell wall remains poorly known and especially its constant modifications during growth as well as environmental changes is not well appreciated. This review focus on the cell wall changes seen between different fungal stages and cell populations with a specific view to explain the resistance to stresses.

Solid-state NMR molecular snapshots of Aspergillus fumigatus cell wall architecture during a conidial morphotype transition

While establishing an invasive infection, the dormant conidia of Aspergillus fumigatus transit through swollen and germinating stages, to form hyphae. During this morphotype transition, the conidial cell wall undergoes dynamic remodeling, which poses challenges to the host immune system and antifungal drugs. However, such cell wall reorganization during conidial germination has not been studied so far. Here, we explored the molecular rearrangement of Aspergillus fumigatus cell wall polysaccharides during different stages of germination. We took advantage of magic-angle spinning NMR to investigate the cell wall polysaccharides, without employing any destructive method for sample preparation. The breaking of dormancy was associated with a significant change in the molar ratio between the major polysaccharides β-1,3-glucan and α-1,3-glucan, while chitin remained equally abundant. The use of various polarization transfers allowed the detection of rigid and mobile polysaccharides; the appearance of mobile galactosaminogalactan was a molecular hallmark of germinating conidia. We also report for the first time highly abundant triglyceride lipids in the mobile matrix of conidial cell walls. Water to polysaccharides polarization transfers revealed an increased surface exposure of glucans during germination, while chitin remained embedded deeper in the cell wall, suggesting a molecular compensation mechanism to keep the cell wall rigidity. We complement the NMR analysis with confocal and atomic force microscopies to explore the role of melanin and RodA hydrophobin on the dormant conidial surface. Exemplified here using Aspergillus fumigatus as a model, our approach provides a powerful tool to decipher the molecular remodeling of fungal cell walls during their morphotype switching.

Aspergillus fumigatus Cell Wall Promotes Apical Airway Epithelial Recruitment of Human Neutrophils.

Aspergillus fumigatus is a ubiquitous fungal pathogen capable of causing multiple pulmonary diseases, including invasive aspergillosis, chronic necrotizing aspergillosis, fungal colonization, and allergic bronchopulmonary aspergillosis. Intact mucociliary barrier function and early airway neutrophil responses are critical for clearing fungal conidia from the host airways prior to establishing disease. Following inhalation, Aspergillus conidia deposit in the small airways, where they are likely to make their initial host encounter with epithelial cells. Challenges in airway infection models have limited the ability to explore early steps in the interactions between A. fumigatus and the human airway epithelium. Here, we use inverted air-liquid interface cultures to demonstrate that the human airway epithelium responds to apical stimulation by A. fumigatus to promote the transepithelial migration of neutrophils from the basolateral membrane surface to the apical airway surface. Promoting epithelial transmigration with Aspergillus required prolonged exposure with live resting conidia. Swollen conidia did not expedite epithelial transmigration. Using A. fumigatus strains containing deletions of genes for cell wall components, we identified that deletion of the hydrophobic rodlet layer or dihydroxynaphthalene-melanin in the conidial cell wall amplified the epithelial transmigration of neutrophils, using primary human airway epithelium. Ultimately, we show that an as-yet-unidentified nonsecreted cell wall protein is required to promote the early epithelial transmigration of human neutrophils into the airspace in response to A. fumigatus Together, these data provide critical insight into the initial epithelial host response to Aspergillus.

In vivo investigation of antimicrobial effects of cinnamaldehyde using immunosuppressed ICR mice with invasive pulmonary aspergillosis

ObjectiveInvasive pulmonary Aspergillus infection has the characteristics of high morbidity, difficult to be treated, poor prognosis and high mortality. This study aims to investigate the effects of cinnamaldehyde on 1,3-β-D-glucans in the pulmonary Aspergillus fumigatus cell wall to provide a basis for developing novel antifungal drugs. MethodsImmunosuppressed ICR mice were intranasally inoculated with 50 µL of A. fumigatus suspension (1 × 107 CFU/mL) and then separated into two groups, for the experimental group cinnamaldehyde was orally administered at 240 mg/kg/d consecutively for 14 d. While for the control group, voriconazole was used to treat the fungus infection. Pulmonary tissues were then extracted for 1,3-β-D-glucans assay and electron microscopy. ResultsThe concentration of 1,3-β-D-glucans was significantly different between the cinnamaldehyde and voriconazole groups, which was (1160.89 ± 364.96) pg/mL and (3885.94 ± 845.45) pg/mL, respectively (P < 0.01). Electron microscopy showed that 2−3 outer layers (1,3-β-D-glucan layer) of A. fumigatus cell wall were damaged and fell off, resulting in serious defect of the cell wall, but the cell membrane was clear and intact. ConclusionCinnamaldehyde has a significant influence on the integrity of 1,3-β-D-glucans in the pulmonary A. fumigatus cell wall, but the cell membrane is unaffected, suggesting that cinnamaldehyde has unique antifungal properties depending on its action against the 1,3-β-D-glucans on the pulmonary A. fumigatus cell wall.

Aspergillus fumigatus exoβ(1-3)glucanases family GH55 are essential for conidial cell wall morphogenesis.

The cell wall of Aspergillus fumigatus is predominantly composed of polysaccharides. The central fibrillar core of the cell wall is composed of a branched β(1-3)glucan, to which the chitin and the galactomannan are covalently bound. Softening of the cell wall is an essential event during fungal morphogenesis, wherein rigid cell wall structures are cleaved by glycosyl hydrolases. In this study, we characterised the role of the glycosyl hydrolase GH55 members in A. fumigatus fungal morphogenesis. We showed that deletion of the six genes of the GH55 family stopped conidial cell wall maturation at the beginning of the development process, leading to abrogation of conidial separation: the shape of conidia became ovoid, and germination was delayed. In conclusion, the reorganisation and structuring of the conidial cell wall mediated by members of the GH55 family is essential for their maturation, normal dissemination, and germination.

The Glycosylphosphatidylinositol-Anchored DFG Family Is Essential for the Insertion of Galactomannan into the β-(1,3)-Glucan-Chitin Core of the Cell Wall of Aspergillus fumigatus.

The fungal cell wall is a complex and dynamic entity essential for the development of fungi. It is composed mainly of polysaccharides that are synthetized by protein complexes. At the cell wall level, enzyme activities are involved in postsynthesis polysaccharide modifications such as cleavage, elongation, branching, and cross-linking. Glycosylphosphatidylinositol (GPI)-anchored proteins have been shown to participate in cell wall biosynthesis and specifically in polysaccharide remodeling. Among these proteins, the DFG family plays an essential role in controlling polar growth in yeast. In the filamentous fungus and opportunistic human pathogen Aspergillus fumigatus, the DFG gene family contains seven orthologous DFG genes among which only six are expressed under in vitro growth conditions. Deletions of single DFG genes revealed that DFG3 plays the most important morphogenetic role in this gene family. A sextuple-deletion mutant resulting from the deletion of all in vitro expressed DFG genes did not contain galactomannan in the cell wall and has severe growth defects. This study has shown that DFG members are absolutely necessary for the insertion of galactomannan into the cell wall of A. fumigatus and that the proper cell wall localization of the galactomannan is essential for correct fungal morphogenesis in A. fumigatusIMPORTANCE The fungal cell wall is a complex and dynamic entity essential for the development of fungi. It is composed mainly of polysaccharides that are synthetized by protein complexes. Enzymes involved in postsynthesis polysaccharide modifications, such as cleavage, elongation, branching, and cross-linking, are essential for fungal life. Here, we investigated in Aspergillus fumigatus the role of the members of the Dfg family, one of the 4 GPI-anchored protein families common to yeast and molds involved in cell wall remodeling. Molecular and biochemical approaches showed that DFG members are required for filamentous growth, conidiation, and cell wall organization and are essential for the life of this fungal pathogen.

β-Glucan Grafted Microcapsule, a Tool for Studying the Immunomodulatory Effect of Microbial Cell Wall Polysaccharides.

β-(1,3)-Glucan is one of the antigenic components of the bacterial as well as fungal cell wall. We designed microcapsules (MCs) ligated with β-(1,3)-glucan, to study its immunomodulatory effect. The MCs were obtained by interfacial polycondensation between diacyl chloride (sebacoyl chloride and terephtaloyl chloride) and diethylenetriamine in organic and aqueous phases, respectively. Planar films were first designed to optimize monomer compositions and to examine the kinetics of film formation. MCs with aqueous fluorescent core were then obtained upon controlled emulsification-polycondensation reactions using optimized monomer compositions and adding fluorescein into the aqueous phase. The selected MC-formulation was grafted with Curdlan, a linear β-(1,3)-glucan from Agrobacterium species or branched β-(1,3)-glucan isolated from the cell wall of Aspergillus fumigatus. These β-(1,3)-glucan grafted MCs were phagocytosed by human monocyte-derived macrophages, and stimulated cytokine secretion. Moreover, the blocking of dectin-1, a β-(1,3)-glucan recognizing receptor, did not completely inhibit the phagocytosis of these β-(1,3)-glucan grafted MCs, suggesting the involvement of other receptors in the recognition and uptake of β-(1,3)-glucan. Overall, grafted MCs are a useful tool for the study of the mechanism of phagocytosis and immunomodulatory effect of the microbial polysaccharides.

Identification of Two Mannosyltransferases Contributing to Biosynthesis of the Fungal-type Galactomannan \u03b1-Core-Mannan Structure in Aspergillus fumigatus

Fungal-type galactomannan (FTGM) is a polysaccharide composed of α-(1 → 2)-/α-(1 → 6)-mannosyl and β-(1 → 5)-/β-(1 → 6)-galactofuranosyl residues located at the outer cell wall of the human pathogenic fungus Aspergillus fumigatus. FTGM contains a linear α-mannan structure called core-mannan composed of 9 or 10 α-(1 → 2)-mannotetraose units jointed by α-(1 → 6)-linkages. However, the enzymes involved in core-mannan biosynthesis remain unknown. We speculated that two putative α-1,2-mannosyltransferase genes in A. fumigatus, Afu5g02740/AFUB_051270 (here termed core-mannan synthase A [CmsA]) and Afu5g12160/AFUB_059750 (CmsB) are involved in FTGM core-mannan biosynthesis. We constructed recombinant proteins for CmsA and detected robust mannosyltransferase activity using the chemically synthesized substrate p-nitrophenyl α-d-mannopyranoside as an acceptor. Analyses of CmsA enzymatic product revealed that CmsA possesses the capacity to transfer a mannopyranoside to the C-2 position of α-mannose. CmsA could also transfer a mannose residue to α-(1 → 2)-mannobiose and α-(1 → 6)-mannobiose and showed a 31-fold higher specific activity toward α-(1 → 6)-mannobiose than toward α-(1 → 2)-mannobiose. Proton nuclear magnetic resonance (1H-NMR) spectroscopy and gel filtration chromatography of isolated FTGM revealed that core-mannan structures were drastically altered and shortened in disruptant A. fumigatus strains ∆cmsA, ∆cmsB, and ∆cmsA∆cmsB. Disruption of cmsA or cmsB resulted in severely repressed hyphal extension, abnormal branching hyphae, formation of a balloon structure in hyphae, and decreased conidia formation. The normal wild type core-mannan structure and developmental phenotype were restored by the complementation of cmsA and cmsB in the corresponding disruptant strains. These findings indicate that both CmsA, an α-1,2-mannosyltransferase, and CmsB, a putative mannosyltransferase, are involved in FTGM biosynthesis.

Effects of Cinnamaldehyde on the Cell Wall of A. fumigatus and Its Application in Treating Mice with Invasive Pulmonary Aspergillosis.

Background The invasive pulmonary aspergillosis is a kind of high incidence of disease with difficulties in treatment, poor prognosis, and high mortality. Objectives The study aimed to reveal the effect of cinnamaldehyde on the fungal cell wall and verify its efficacy on invasive pulmonary aspergillosis on immunosuppressed Institute of Cancer Research mice (ICR mice). Methods ICR mice were given cyclophosphamide 200 mg.kg−1. d−1 by intraperitoneal injection for 2 days. On the 4th day, the mice were given 50 μL of Aspergillosis fumigatus spore (107colony form unit CFU/mL) by intranasal injection to establish immunosuppressive animal models with invasive Aspergillosis fumigatus infection. Then the mice in treatment group orally administered cinnamaldehyde for 14 consecutive days, while voriconazole was given to the mice in the positive control group. Results The clearance rate of pulmonary fungi, cure rate, and reduction of 1,3-β-D-glucans in treatment group were 80.00%, 80.00%, and 81.00%, respectively while in positive control group they were 67.00%, 60.00%, and 62.00%, respectively. There were significant differences in the results between two groups as mentioned above (P<0.05). Electron microscopy showed that, in treatment group, the cell wall of Aspergillus fumigatus was dissolved and detached and the cell surface was incomplete. There were edema, degeneration, and necrosis in nucleus and organelle, which lead to cellular necrocytosis. The cytomembrane of Aspergillus fumigatus was intact, clear, and complete, whereas the cytomembrane in the positive control group disappeared. The hyphal morphology of Aspergillus fumigatus was deformed, but the cell wall was intact. Conclusion Cinnamaldehyde has a good curative effect in the treatment of invasive pulmonary aspergillus infection in immunodeficient mice. It mainly affects the synthesis of 1,3-β-D-glucans from the cytoderm of Aspergillus fumigatus but does not affect cell wall. It would potentially be an effective and novel drug for targeted treatment of Aspergillus fumigatus deep infection.

Study of the Carbohydrate Specificity of Antibodies Against Aspergillus fumigatus Using the Library of Synthetic Mycoantigens

The carbohydrate specificity of antibodies obtained by immunizing laboratory animals with immunogens prepared from the Aspergillus fumigatus cell wall was analyzed using the library of biotinylated synthetic oligosaccharides. It has been shown that the main part of antipolysaccharide antibodies recognizes galactomannan in the studied immunogens with preference to its fragments containing more than two β-(1→5)-linked galactofuranosyl units. The data obtained can form the basis for the development of enzyme immunoassay for the detection of this dangerous fungal pathogen.

Aspergillus fumigatus Cell Wall α-(1,3)-Glucan Stimulates Regulatory T-Cell Polarization by Inducing PD-L1 Expression on Human Dendritic Cells.

Human dendritic cell (DC) response to α-(1,3)-glucan polysaccharide of Aspergillus fumigatus and ensuing CD4+ T-cell polarization are poorly characterized. α-(1,3)-Glucan was isolated from A. fumigatus conidia and mycelia cell wall. For the analysis of polarization, DCs and autologous naive CD4+ T cells were cocultured. Phenotype of immune cells was analyzed by flow cytometry, and cytokines by enzyme-linked immunosorbent assay (ELISA). Blocking antibodies were used to dissect the role of Toll-like receptor 2 (TLR2) and programmed death-ligand 1 (PD-L1) in regulating α-(1,3)-glucan-mediated DC activation and T-cell responses. DCs from TLR2-deficient mice were additionally used to consolidate the findings. α-(1,3)-Glucan induced the maturation of DCs and was dependent in part on TLR2. "α-(1,3)-Glucan-educated" DCs stimulated the activation of naive T cells and polarized a subset of these cells into CD4+CD25+FoxP3+ regulatory T cells (Tregs). Mechanistically, Treg stimulation by α-(1,3)-glucan was dependent on the PD-L1 pathway that negatively regulated interferon-gamma (IFN-γ) secretion. Short α-(1,3)-oligosaccharides lacked the capacity to induce maturation of DCs but significantly blocked α-(1,3)-glucan-induced Treg polarization. PD-L1 dictates the balance between Treg and IFN-γ responses induced by α-(1,3)-glucan. Our data provide a rationale for the exploitation of immunotherapeutic approaches that target PD-1-PD-L1 to enhance protective immune responses to A. fumigatus infections.

The Cell Wall of the Human Fungal Pathogen Aspergillus fumigatus: Biosynthesis, Organization, Immune Response, and Virulence.

More than 90% of the cell wall of the filamentous fungus Aspergillus fumigatus comprises polysaccharides. Biosynthesis of the cell wall polysaccharides is under the control of three types of enzymes: transmembrane synthases, which are anchored to the plasma membrane and use nucleotide sugars as substrates, and cell wall-associated transglycosidases and glycosyl hydrolases, which are responsible for remodeling the de novo synthesized polysaccharides and establishing the three-dimensional structure of the cell wall. For years, the cell wall was considered an inert exoskeleton of the fungal cell. The cell wall is now recognized as a living organelle, since the composition and cellular localization of the different constitutive cell wall components (especially of the outer layers) vary when the fungus senses changes in the external environment. The cell wall plays a major role during infection. The recognition of the fungal cell wall by the host is essential in the initiation of the immune response. The interactions between the different pattern-recognition receptors (PRRs) and cell wall pathogen-associated molecular patterns (PAMPs) orientate the host response toward either fungal death or growth, which would then lead to disease development. Understanding the molecular determinants of the interplay between the cell wall and host immunity is fundamental to combatting Aspergillus diseases.

Recent advances in the understanding of the Aspergillus fumigatus cell wall.

Over the past several decades, research on the synthesis and organization of the cell wall polysaccharides of Aspergillus fumigatus has expanded our knowledge of this important fungal structure. Besides protecting the fungus from environmental stresses and maintaining structural integrity of the organism, the cell wall is also the primary site for interaction with host tissues during infection. Cell wall polysaccharides are important ligands for the recognition of fungi by the innate immune system and they can mediate potent immunomodulatory effects. The synthesis of cell wall polysaccharides is a complicated process that requires coordinated regulation of many biosynthetic and metabolic pathways. Continuous synthesis and remodeling of the polysaccharides of the cell wall is essential for the survival of the fungus during development, reproduction, colonization and invasion. As these polysaccharides are absent from the human host, these biosynthetic pathways are attractive targets for antifungal development. In this review, we present recent advances in our understanding of Aspergillus fumigatus cell wall polysaccharides, including the emerging role of cell wall polysaccharides in the host-pathogen interaction.

The Aspergillus fumigatus cell wall integrity signaling pathway: drug target, compensatory pathways, and virulence.

Aspergillus fumigatus is the most important airborne fungal pathogen, causing severe infections with invasive growth in immunocompromised patients. The fungal cell wall (CW) prevents the cell from lysing and protects the fungus against environmental stress conditions. Because it is absent in humans and because of its essentiality, the fungal CW is a promising target for antifungal drugs. Nowadays, compounds acting on the CW, i.e., echinocandin derivatives, are used to treat A. fumigatus infections. However, studies demonstrating the clinical effectiveness of echinocandins in comparison with antifungals currently recommended for first-line treatment of invasive aspergillosis are still lacking. Therefore, it is important to elucidate CW biosynthesis pathways and their signal transduction cascades, which potentially compensate the inhibition caused by CW- perturbing compounds. Like in other fungi, the central core of the cell wall integrity (CWI) signaling pathway in A. fumigatus is composed of three mitogen activated protein kinases. Deletion of these genes resulted in severely enhanced sensitivity of the mutants against CW-disturbing compounds and in drastic alterations of the fungal morphology. Additionally, several cross-talk interactions between the CWI pathways and other signaling pathways are emerging, raising the question about their role in the CW compensatory mechanisms. In this review we focused on recent advances in understanding the CWI signaling pathway in A. fumigatus and its role during drug stress response and virulence.

Synthesis of a pentasaccharide and neoglycoconjugates related to fungal α-(1→3)-glucan and their use in the generation of antibodies to trace Aspergillus fumigatus cell wall.

3-Aminopropyl α-(1→3)-pentaglucoside, a fragment of α-(1→3)-glucan of the cell wall of Aspergillus fumigatus, has been synthesized in a blockwise approach. The application of mono- and disaccharide N-phenyltrifluoroacetimidates bearing a stereodirecting 6-O-benzoyl group was essential for stereoselective α-glucosylations. In the products, p-methoxyphenyl and levulinoyl groups served as orthogonal protecting groups for the anomeric position and 3-OH group, respectively. Their removal from shared blocks led to donors and acceptors that were used for the synthesis of pentasaccharides. Coupling of free α-(1→3)-pentaglucoside with biotin and bovine serum albumin (BSA) gave glycoconjugate tools for mycological studies. Immunization of mice with the BSA conjugate induced the generation of antibodies that recognize α-(1→3)-glucan on A. fumigatus cell wall and distinguish its morphotypes. This discovery represents a first step to the development of a diagnostic test system and a vaccine to detect and fight this life-threatening pathogen.

Synthesis of a Pentasaccharide and Neoglycoconjugates Related to Fungal α-(1→3)-Glucan and Their Use in the Generation of Antibodies to TraceAspergillus fumigatusCell Wall

Invited for the cover of this issue is the group of researchers from N.D. Zelinsky Institute of the Russian Academy of Sciences and the Pasteur Institute, Paris. The coverpicture illustrates our research: the production of antibodies against Aspergillus fumigatus cell wall α-(1→3)-glucan using chemically prepared pentasaccharide fragment of α-(1→3)-glucan conjugated with a carrier protein. Read the full text of the article at 10.1002/chem.201404770.

Aspergillus cell wall and biofilm.

The fungal cell is surrounded by a cell wall that acts as a sieve and a reservoir for effector molecules that play an active role during infection. This cell wall is essential for fungal growth as well as for resisting host defense mechanisms. The Aspergillus fumigatus cell wall is almost exclusively composed of polysaccharides. The fibrillar core is composed of a branched β-(1,3)-glucan to which chitin, β-(1,3)-/β-(1,4)-glucan, and galactomannan are covalently bound. The alkali-soluble amorphous fraction is mainly composed of α-(1,3)-glucan that has adhesive property and stabilizes the cell wall. Although the same polysaccharides are found in the cell wall of different A. fumigatus morphotypes (conidia and hyphae), their concentration and localization are different. Conidial (the morphotype that mainly enters host respiratory system) cell wall is covered by an outer layer of rodlets and melanin, which confers hydrophobic properties and imparts immunological inertness. In contrast, outer layer of the hypha contains galactosaminogalactan, recently identified as an A. fumigatus virulence factor. The hypha grows either as a network of agglutinated and hydrophobic mass (called mycelium) embedded in an extracellular matrix (ECM) rich in polysaccharides, hydrophobin, and melanin or segregated without ECM.

Overlapping and Distinct Roles of Aspergillus fumigatus UDP-glucose 4-Epimerases in Galactose Metabolism and the Synthesis of Galactose-containing Cell Wall Polysaccharides

The cell wall of Aspergillus fumigatus contains two galactose-containing polysaccharides, galactomannan and galactosaminogalactan, whose biosynthetic pathways are not well understood. The A. fumigatus genome contains three genes encoding putative UDP-glucose 4-epimerases, uge3, uge4, and uge5. We undertook this study to elucidate the function of these epimerases. We found that uge4 is minimally expressed and is not required for the synthesis of galactose-containing exopolysaccharides or galactose metabolism. Uge5 is the dominant UDP-glucose 4-epimerase in A. fumigatus and is essential for normal growth in galactose-based medium. Uge5 is required for synthesis of the galactofuranose (Galf) component of galactomannan and contributes galactose to the synthesis of galactosaminogalactan. Uge3 can mediate production of both UDP-galactose and UDP-N-acetylgalactosamine (GalNAc) and is required for the production of galactosaminogalactan but not galactomannan. In the absence of Uge5, Uge3 activity is sufficient for growth on galactose and the synthesis of galactosaminogalactan containing lower levels of galactose but not the synthesis of Galf. A double deletion of uge5 and uge3 blocked growth on galactose and synthesis of both Galf and galactosaminogalactan. This study is the first survey of glucose epimerases in A. fumigatus and contributes to our understanding of the role of these enzymes in metabolism and cell wall synthesis.