Biology subjects

Naglik, J. R.

Publications and source records attributed to Naglik, J. R..

4 recordsLinked to original sources

bioRxiv2023

EGR1 regulates oral epithelial cell responses to Candida albicans via the EGFR- ERK1/2 pathway

Candida albicans is a fungal pathobiont colonising mucosal surfaces of the human body, including the oral cavity. Under certain predisposing conditions, C. albicans invades mucosal tissues activating EGFR-MAPK signalling pathways in epithelial cells via the action of its peptide toxin candidalysin. However, our knowledge of the epithelial mechanisms involved during C. albicans colonisation is rudimentary. Here, we describe the role of the transcription factor early growth response protein 1 (EGR1) in human oral epithelial cells (OECs) in response to C. albicans. EGR1 expression increases in OECs when exposed to C. albicans independently of fungal viability, morphology, or candidalysin release, suggesting EGR1 is involved in the fundamental recognition of C. albicans, rather than in response to invasion or pathogenesis. Upregulation of EGR1 is mediated by EGFR via Raf1, ERK1/2 and NF-{kappa}B signalling but not PI3K/mTOR signalling. Notably, EGR1 mRNA silencing impacts on anti-C. albicans immunity, reducing GM-CSF, IL-1 and IL-1{beta} release, and increasing IL-6 and IL-8 production. These findings identify an important role for EGR1 in priming epithelial cells to respond to subsequent invasive infection by C. albicans and elucidate the regulation circuit of this transcription factor after contact.

immunology↗

bioRxiv2022

Candida albicans promotes neutrophil extracellular trap formation and leukotoxic hypercitrullination via the peptide toxin candidalysin.

The cytolytic peptide toxin candidalysin is secreted by the invasive, hyphal form of the human fungal pathogen, Candida albicans. Candidalysin is essential for inducing host cell damage during mucosal and systemic C. albicans infections, resulting in neutrophil recruitment. Neutrophil influx to C. albicans-infected tissue is critical for limiting fungal growth and preventing the fungal dissemination. Here, we demonstrate that candidalysin secreted by hyphae promotes the stimulation of neutrophil extracellular traps (NETs), while synthetic candidalysin triggers a distinct mechanism for NET-like structures (NLS), which are more compact and less fibrous than canonical NETs. Candidalysin activates NADPH oxidase and calcium influx, with both processes contributing to morphological changes in neutrophils resulting in NLS formation. NLS are induced by leukotoxic hypercitrullination, which is governed by protein arginine deaminase 4 activation via calcium influx and initiation of intracellular signalling events. However, activation of signalling by candidalysin does not suffice to trigger downstream events essential for NET formation, as demonstrated by lack of lamin A/C phosphorylation, an event required for activation of cyclin-dependent kinases that are crucial for NET release. Interestingly, exposure to candidalysin does not immediately restrict the capability of neutrophils to produce reactive oxygen species (ROS), nor to phagocytose particles. Instead, candidalysin triggers ROS production, calcium influx and subsequent activation of downstream signalling that drive morphological alteration and the formation of NLS in a dose- and time-dependent manner. Notably, candidalysin-triggered NLS demonstrate anti-Candida activity, which is resistant to nuclease treatment and dependent on the deprivation of Zn2+. This study reveals that C. albicans hyphae releasing candidalysin concurrently trigger canonical NETs and NLS, which together form a fibrous sticky network that entangles C. albicans hyphae and inhibits their growth. Importantly, this explains discrepancies of previous studies demonstrating that neutrophil-derived extracellular chromatin structures triggered by C. albicans can be both dependent and independent of ROS. Our data also demonstrate that while candidalysin hampers neutrophil function, the toxin also increases the capability of neutrophils to entangle hyphae and to restrict their growth, reflecting the importance of human neutrophils in controlling the dissemination of C. albicans.

immunology↗

bioRxiv2022

EGFR-MAPK adaptor proteins mediate the epithelial response to Candida albicans via the cytolytic peptide toxin, candidalysin

Candida albicans (C. albicans) is a dimorphic human fungal pathogen that can cause severe oropharyngeal candidiasis (OPC, oral thrush) in susceptible hosts. During invasive infection, C. albicans hyphae invade oral epithelial cells (OECs) and secrete candidalysin, a pore-forming cytolytic peptide that is required for fungal pathogenesis at mucosal surfaces. Candidalysin induces cell damage and activates multiple MAPK-based innate signaling events that collectively drive the production of downstream inflammatory mediators. The activities of candidalysin are also dependent on the epidermal growth factor receptor (EGFR), but how these signals are integrated is undefined. Here, we identified five essential adaptor proteins as key mediators of the epithelial response to C. albicans infection on cultured OECs, including growth factor receptor bound protein 2 (Grb2), Grb2-associated-binding protein 1 (Gab1), Src homology and collagen (Shc), SH2 containing protein tyrosine phosphatase-2 (Shp2) and casitas B-lineage lymphoma (c-Cbl). All these signaling effectors were inducibly phosphorylated in response to C. albicans, in a candidalysin-dependent mechanism but additionally required EGFR phosphorylation, matrix metalloproteinases (MMPs) and cellular calcium flux. Of these, Gab1, Grb2 and Shp2 were the dominant drivers of ERK1/2 signaling and production of downstream cytokines. Together, these results identify the key adaptor proteins that drive EGFR signaling mechanisms, which determine oral epithelial responses to C. albicans.

microbiology↗

bioRxiv2021

Aspergillus fumigatus drives tissue damage via iterative assaults upon mucosal integrity and immune homeostasis

The human lung is constantly exposed to Aspergillus fumigatus spores, the most prevalent worldwide cause of fungal respiratory disease. Pulmonary tissue damage is a unifying feature of Aspergillus-related diseases; however, the mechanistic basis of damage is not understood. In the lungs of susceptible hosts A. fumigatus undergoes an obligatory morphological switch involving spore germination and hyphal growth. We modelled A. fumigatus infection in cultured A549 human pneumocytes, capturing phosphoactivation status of five host signalling pathways, nuclear translocation & DNA binding of eight host transcription factors, and expression of nine host response proteins over six time points encompassing exposures to live fungus and the secretome thereof. The resulting dataset, comprised of more than 1000 data points, reveals that pneumocytes mount differential responses to A. fumigatus spores, hyphae and soluble secreted products via the NF-kB, JNK, and JNK + p38 pathways respectively. Importantly, via selective degradation of host pro-inflammatory (IL-6 and IL-8) cytokines and growth factors (FGF-2), fungal secreted products reorchestrate the host response to fungal challenge as well as driving multiparametric epithelial damage, culminating in cytolysis. Dysregulation of NF-kB signalling, involving iterative stimulation of canonical and non-canonical signalling, was identified as a significant feature of host damage both in vitro and in a mouse model of invasive aspergillosis. Our data demonstrate that composite tissue damage results from iterative exposures to different fungal morphotypes and secreted products and suggest that modulation of host responses to fungal challenge might represent a unified strategy for therapeutic control of pathologically distinct types of Aspergillus-related disease. IMPORTANCEPulmonary aspergillosis is a spectrum of diseases caused primarily by Aspergillus fumigatus. This fungus is ubiquitous in the environment and grows as a mold, which harbors and disperses spores into the environment. Like other airborne pathogens, the lung mucosa is the first point of contact with the fungus post inhalation. The outcome and severity of disease depends on the host-fungal interaction at the lung interface. We studied how the human lung interacts with spore, germ tube and hyphae growth forms to understand the sequence and dynamics of the early events, which are critical drivers of disease development and progression. Our work is significant in identifying, in response to fungal secreted products, non-canonical NF-kB activation via RelB as being a driving factor in fungus-mediated lung damage. This process could be modulated therapeutically to protect the integrity of infected lung mucosae.

microbiology↗