Biology subjects

Phan, Q. T.

Publications and source records attributed to Phan, Q. T..

8 recordsLinked to original sources

bioRxiv2025

The HosA histone deacetylase regulates stress resistance, host cell interactions, and virulence in Aspergillus fumigatus

The capacity of Aspergillus fumigatus to cause invasive pulmonary aspergillosis depends on its ability to adapt to dynamic and stressful microenvironments within the host. Epigenetic regulation, including histone deacetylation, plays a critical role in fungal adaptation to stress. Here, we investigated the role of the class I histone deacetylase (HDAC) HosA in A. fumigatus stress resistance, host cell interactions, and virulence. A {Delta}hosA mutant had increased susceptibility to intracellular oxidant stress induced by menadione. It also had impaired capacity to invade and damage two pulmonary epithelial cell lines in vitro. In a corticosteroid-immunosuppressed mouse model of invasive aspergillosis, mice infected with the {Delta}hosA mutant survived significantly longer than those infected with the wild-type strain, despite having similar pulmonary fungal burden. The {Delta}hosA mutant also induced a weaker inflammatory response than the wild-type strain. Transcriptomic analysis revealed that HosA regulates genes involved in secondary metabolite biosynthesis and energy metabolism, functioning as both an activator and repressor of distinct gene sets. Collectively, these results indicate that HosA is a key epigenetic regulator that governs A. fumigatus interactions with host cells and virulence during invasive pulmonary aspergillosis. ImportanceEpigenetic modifications in A. fumigatus can be induced by environmental changes and stresses such as those induced by interaction with host cells. HosA, a class I histone deacetylase, has been shown to play a key role in regulating secondary metabolism in several Aspergillus species, but its function in A. fumigatus was previously unknown. We found that deletion of hosA increased susceptibility to intracellular, but not extracellular, oxidative stress. The {Delta}hosA mutant also exhibited significantly reduced pulmonary epithelial cell invasion and host cell damage, as well as attenuated virulence in immunosuppressed mice. Together, these findings indicate that HosA functions as a key epigenetic regulator that governs stress resistance, secondary metabolism, and fungal-host interactions. Defining the functions of HosA could provide critical insight into the epigenetic mechanisms that control fungal pathogenicity and may reveal a potential therapeutic target for invasive aspergillosis.

microbiology↗

bioRxiv2024

Epidermal Growth Factor Receptor Signaling Governs the Host Inflammatory Response to Invasive Aspergillosis

The epidermal growth factor receptor (EGFR) has been identified as an epithelial cell receptor for Mucorales fungi and Candida albicans. Blocking EGFR with small molecule inhibitors reduces disease severity in mouse models of mucormycosis and oropharyngeal candidiasis. In contrast, cases of invasive aspergillosis have been reported in cancer patients who were treated with EGFR inhibitors, suggesting that EGFR signaling may play a protective role in the host defense against this infection. Here, we analyzed transcriptomic data from the lungs of mice with invasive aspergillosis and found evidence that Aspergillus fumigatus infection activates multiple genes that are predicted to function in the EGFR signaling pathway. We also found that A. fumigatus infection activates EGFR in both a human small airway epithelial (HSAE) cell line and in the lungs of immunosuppressed mice. EGFR signaling in HSAE cells is required for maximal endocytosis of A. fumigatus and for fungal-induced proinflammatory cytokine and chemokine production. In a corticosteroid immunosuppressed mouse model of invasive pulmonary aspergillosis, inhibition of EGFR with gefitinib decreased whole lung chemokine levels and reduced accumulation of phagocytes in the lung, leading to a decrease in fungal killing, an increase in pulmonary fungal burden, and accelerated mortality. Thus, EGFR signaling is required for pulmonary epithelial cells to orchestrate the host innate immune defense against invasive aspergillosis in immunosuppressed hosts. ImportanceWhen A. fumigatus infects the lungs, it invades epithelial cells that line the airways. During this process, the fungus interacts with epithelial cell receptors. This interaction stimulates epithelial cells to endocytose the fungus. It also induces these cells to secret proinflammatory cytokines and chemokines that recruit phagocytes to the site of infection where they can kill the fungus. Here, we show that in small airway epithelial cells, the epidermal growth factor receptor (EGFR) acts a sensor for A. fumigatus that triggers the production of chemokines in response to fungal infection. In corticosteroid-immunosuppressed mice, blocking EGFR with the kinase inhibitor, gefitinib reduces chemokine production in the lungs. This leads to decreased accumulation of neutrophils and dendritic cell in the lungs, reduced A. fumigatus killing, and increased mortality. These results provide a potential explanation as to why some cancer patients who are treated with EGFR inhibitors develop invasive aspergillosis.

microbiology↗

bioRxiv2024

A genome-scale screen identifies sulfated glycosaminoglycans as pivotal in epithelial cell damage by Candida albicans

Candidalysin is a cytolytic peptide produced by the opportunistic fungal pathogen Candida albicans. This peptide is a key virulence factor in mouse models of mucosal and hematogenously disseminated candidiasis. Despite intense interest in the role of candidalysin in C. albicans pathogenicity, its host cell targets have remained elusive. To fill this knowledge gap, we performed a genome-wide loss-of-function CRISPR screen in a human oral epithelial cell line to identify specific host factors required for susceptibility to candidalysin-induced cellular damage. Among the top hits were XYLT2, B3GALT6 and B3GAT3, genes that function in glycosaminoglycan (GAG) biosynthesis. Deletion of these genes led to the absence of GAGs such as heparan sulfate on the epithelial cell surface and increased resistance to damage induced by both candidalysin and live C. albicans. Biophysical analyses including surface plasmon resonance and atomic force and electron microscopy indicated that candidalysin physically binds to sulfated GAGs, facilitating its oligomerization or enrichment on the host cell surface. The addition of exogenous sulfated GAGs or the GAG analogue dextran sulfate protected cells against candidalysin-induced damage. Dextran sulfate, but not non-sulfated dextran, also inhibited epithelial cell endocytosis of C. albicans and fungal-induced epithelial cell cytokine and chemokine production. In a murine model of vulvovaginal candidiasis, topical dextran sulfate administration reduced host tissue damage and decreased intravaginal IL-1{beta} and neutrophil levels. Collectively, these data indicate that GAGs are epithelial cell targets of candidalysin and can be used therapeutically to protect cells from candidalysin-induced damage.

microbiology↗

bioRxiv2023

Use of a human small airway epithelial cell line to study the interactions of Aspergillus fumigatus with pulmonary epithelial cells

During the initiation of invasive aspergillosis, inhaled Aspergillus fumigatus conidia are deposited on the epithelial cells lining the bronchi, terminal bronchioles, and alveoli. While the interactions of A. fumigatus with bronchial and type II alveolar cell lines have been investigated in vitro, little is known about the interactions of this fungus with terminal bronchiolar epithelial cells. We compared the interactions of A. fumigatus with the A549 type II alveolar epithelial cell line and the HSAEC1-KT human small airway epithelial (HSAE) cell line. We found that A. fumigatus conidia were poorly endocytosed by A549 cells, but avidly endocytosed by HSAE cells. A. fumigatus germlings invaded both cell types by induced endocytosis, but not by active penetration. A549 cell endocytosis of A. fumigatus was independent of fungal viability, more dependent on host microfilaments than microtubules, and induced by A. fumigatus CalA interacting with host cell integrin 5{beta}1. By contrast, HSAE cell endocytosis required fungal viability, was more dependent on microtubules than microfilaments, and did not require CalA or integrin 5{beta}1. HSAE cells were more susceptible than A549 cells to damage caused by direct contact with killed A. fumigatus germlings and by secreted fungal products. In response to A. fumigatus infection, A549 cells secreted a broader profile of cytokines and chemokines than HSAE cells. Taken together, these results demonstrate that studies of HSAE cells provide complementary data to A549 cells and thus represent a useful model for probing the interactions of A. fumigatus with bronchiolar epithelial cells in vitro. ImportanceDuring the initiation of invasive aspergillosis, Aspergillus fumigatus invades, damages, and stimulates the epithelial cells that line the airways and alveoli. Previous studies of A. fumigatus- epithelial cell interactions in vitro have used either large airway epithelial cell lines or the A549 type II alveolar epithelial cell line. The interactions of fungi with terminal bronchiolar epithelial cells have not been investigated. Here, we compared the interactions of A. fumigatus with A549 cells and the Tert-immortalized human small airway epithelial HSAEC1-KT (HSAE) cell line. We discovered that A. fumigatus invades and damages these two cell lines by distinct mechanisms. Also, the proinflammatory responses of the cell lines to A. fumigatus are different. These results provide insight into how A. fumigatus interacts with different types of epithelial cells during invasive aspergillosis and demonstrate that HSAE cells are useful in vitro model for investigating the interactions of this fungus with bronchiolar epithelial cells.

microbiology↗

bioRxiv2023

Candida albicans stimulates the formation of a multi-receptor complex that mediates epithelial cell invasion during oropharyngeal infection

Fungal invasion of the oral epithelium is central to the pathogenesis of oropharyngeal candidiasis (OPC). Candida albicans invades the oral epithelium by receptor-induced endocytosis but this process is incompletely understood. We found that C. albicans infection of oral epithelial cells induces c-Met to form a multi-protein complex with E-cadherin and the epidermal growth factor receptor (EGFR). E-cadherin is necessary for C. albicans to activate both c-Met and EGFR and to induce the endocytosis of C. albicans. Proteomics analysis revealed that c-Met interacts with C. albicans Hyr1, Als3 and Ssa1. Both Hyr1 and Als3 were required for C. albicans stimulation of c-Met and EGFR in oral epithelial cells in vitro and for full virulence during OPC in mice. Treating mice with small molecule inhibitors of c-Met and EGFR ameliorated OPC, demonstrating the potential therapeutic efficacy of blocking these host receptors for C. albicans. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=161 HEIGHT=200 SRC="FIGDIR/small/529756v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1448372org.highwire.dtl.DTLVardef@111b411org.highwire.dtl.DTLVardef@144f3b8org.highwire.dtl.DTLVardef@1dcf548_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIc-Met is an oral epithelial cell receptor for Candida albicans C_LIO_LIC. albicans infection causes c-Met and the epidermal growth factor receptor (EGFR) to form a complex with E-cadherin, which is required for c-Met and EGFR function C_LIO_LIC. albicans Hyr1 and Als3 interact with c-Met and EGFR, inducing oral epithelial cell endocytosis and virulence during oropharyngeal candidiasis C_LIO_LIDual blockade of c-Met and EGFR ameliorates oropharyngeal candidiasis C_LI

microbiology↗

bioRxiv2021

Identification of Serum Bridging Molecules that Mediate Human Endothelial Cell Invasion by Candida species

During hematogenously disseminated candidiasis, blood borne fungi must invade the endothelial cells that line the blood vessels to infect the deep tissues. Although Candida albicans, which forms hyphae, readily invades endothelial cells, other medically important species of Candida are poorly invasive in standard in vitro assays. Here, we show that Candida glabrata, Candida tropicalis, Candida parapsilosis, and Candida krusei can bind to vitronectin and high molecular weight kininogen present in human serum. Acting as bridging molecules, vitronectin and kininogen bind to v integrins and the globular C1q receptor (gC1qR), inducing human endothelial cells to endocytose the fungus. This mechanism of endothelial cell invasion is poorly supported by mouse endothelial cells, but can be restored when mouse endothelial cells are engineered to express human gC1qR or v integrin. Overall, these data indicate that bridging molecule-mediated endocytosis is a common pathogenic strategy used by many medically important Candida spp. to invade human vascular endothelial cells. SignificanceThe invasion of vascular endothelial cells is a key step in the pathogenesis of hematogenously disseminated candidiasis. How species of Candida other than C. albicans invade endothelial cells is poorly understood because these fungi are weakly invasive in serum-free media. Here, we demonstrate that C. glabrata and other Candida spp. bind to the serum proteins kininogen and vitronectin, which act as bridging molecules and mediate the adherence and endocytosis of the organisms by endothelial cells. These serum proteins induce endocytosis when they interact with the globular C1q receptor and v integrins on human, but not mouse endothelial cells. Thus, bridging molecule-mediated endocytosis is a common mechanism by which medically important Candida spp. invade human endothelial cells.

microbiology↗

bioRxiv2021

The globular C1q receptor is required for epidermal growth factor receptor signaling during Candida albicans infection

During oropharyngeal candidiasis, Candida albicans activates the epidermal growth factor receptor (EGFR), which induces oral epithelial cells to both endocytose the fungus and synthesize proinflammatory mediators that orchestrate the host immune response. To elucidate the signaling pathways that are stimulated when C. albicans interacts with EGFR, we analyzed the proteins that associate with EGFR when C. albicans infects human oral epithelial cells. We identified 1214 proteins that were associated with EGFR in C. albicans-infected cells. We investigated the function of seven of these proteins that either showed increased association with EGFR in response to C. albicans or that mediated the interaction of other microbial pathogens with epithelial cells. Among these proteins, EGFR was found to associate with WW domain-binding protein 2, toll-interacting protein, interferon-induced transmembrane protein 3, and the globular C1q receptor (gC1qR) in viable epithelial cells. Each of these proteins was required for maximal endocytosis of C. albicans and they all regulated fungal-induced production of IL-1{beta} and/or IL-8, either positively or negatively. gC1qR functioned as a key coreceptor with EGFR. Interacting with the C. albicans Als3 invasin, gC1qR was required for the fungus to stimulate both EGFR and the ephrin type-A receptor 2. The combination of gC1qR and EGFR was necessary for maximal endocytosis of C. albicans and secretion of IL-1{beta}, IL-8, and GM-CSF. Thus, this work provides an atlas of proteins that associate with EGFR and identifies several that play a central role in the response of human oral epithelial cells to C. albicans infection. IMPORTANCEOral epithelial cells play a key role in the pathogenesis of oropharyngeal candidiasis. In addition to being target host cells for C. albicans adherence and invasion, they secrete proinflammatory cytokines and chemokines that recruit T cells and activated phagocytes to foci of infection. It is known that C. albicans activates EGFR on oral epithelial cells, which induces these cells to endocytose the organism and stimulates them to secrete proinflammatory mediators. To elucidate the EGFR signaling pathways that govern these responses, we analyzed the epithelial cell proteins that associate with EGFR in C. albicans-infected epithelial cells. We identified four proteins that physically associate with EGFR and that regulate different aspects of the epithelial response to C. albicans. One of these is gC1qR, which is required for C. albicans to activate EGFR, induce endocytosis, and stimulate the secretion of proinflammatory mediators, indicating that gC1qR functions as a key co-receptor with EGFR.

microbiology↗

bioRxiv2020

Determining Aspergillus fumigatus transcription factor expression and function during invasion of the mammalian lung

To gain a better understanding of the transcriptional response of Aspergillus fumigatus during invasive pulmonary infection, we used a NanoString nCounter to assess the transcript levels of 467 A. fumigatus genes during growth in the lungs of immunosuppressed mice. These genes included ones known to respond to diverse environmental conditions and those encoding most transcription factors in the A. fumigatus genome. We found that invasive growth in vivo induces a unique transcriptional profile as the organism responds to nutrient limitation and attack by host phagocytes. This in vivo transcriptional response is largely mimicked by in vitro growth in Aspergillus minimal medium that is deficient in nitrogen, iron, and/or zinc. From the transcriptional profiling data, we selected 9 transcription factor genes that were either highly expressed or strongly up-regulated during in vivo growth. Deletion mutants were constructed for each of these genes and assessed for virulence in mice. Two transcription factor genes were found to be required for maximal virulence. One was rlmA, which governs the ability of the organism to proliferate in the lung. The other was ace1, which regulates of the expression of multiple secondary metabolite gene clusters and mycotoxin genes independently of laeA. Using deletion and overexpression mutants, we determined that the attenuated virulence of the {Delta}ace1 mutant is due to decreased expression aspf1, which specifies a ribotoxin, but is not mediated by reduced expression of the fumigaclavine gene cluster or the fumagillin-pseruotin supercluster. Thus, in vivo transcriptional profiling focused on transcription factors genes provides a facile approach to identifying novel virulence regulators. Author summaryAlthough A. fumigatus causes the majority of cases of invasive aspergillosis, the function of most of the genes in its genome remains unknown. To identify genes encoding transcription factors that may be important for virulence, we used a NanoString nCounter to measure the mRNA levels of A. fumigatus transcription factor genes in the lungs of mice with invasive aspergillosis. The transcriptional profiling data indicate that the organism is exposed to nutrient limitation and stress during growth in the lungs, and that it responds by up-regulating genes that encode mycotoxins and secondary metabolites. In vitro, this response was most closely mimicked by growth in medium that was deficient in nitrogen, iron and/or zinc. Using the transcriptional profiling data, we identified two transcription factors that govern A. fumigatus virulence. These were RlmA, which is governs proliferation in the lung and Ace1, which controls the production of mycotoxins and secondary metabolites.

microbiology↗