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Airway inflammation and remodelling are important pathophysiologic features in asthma and other respiratory conditions. An intact epithelial cell layer is crucial to maintain lung homoeostasis, and this depends on intercellular adhesion, whilst damaged respiratory epithelium is the primary instigator of airway inflammation. The Coxsackievirus Adenovirus Receptor (CAR) is highly expressed in the epithelium where it modulates cell-cell adhesion stability and facilitates immune cell transepithelial migration. However, the contribution of CAR to lung inflammation remains unclear. Here we investigate the mechanistic contribution of CAR in mediating responses to the common aeroallergen, House Dust Mite (HDM). We demonstrate that administration of HDM in mice lacking CAR in the respiratory epithelium leads to loss of peri-bronchial inflammatory cell infiltration, fewer goblet-cells and decreased pro-inflammatory cytokine release. In vitro analysis in human lung epithelial cells confirms that loss of CAR leads to reduced HDM-dependent inflammatory cytokine release and neutrophil migration. Epithelial CAR depletion also promoted smooth muscle cell proliferation mediated by GSK3β and TGF-β, basal matrix production and airway hyperresponsiveness. Our data demonstrate that CAR coordinates lung inflammation through a dual function in leucocyte recruitment and tissue remodelling and may represent an important target for future therapeutic development in inflammatory lung diseases. The epithelial protein Coxsackievirus Adenovirus Receptor (CAR) is a virus receptor but may have other functions. Here the authors show that deletion of CAR in mice leads to reduced house dust mite-induced lung inflammation, reduced neutrophil accumulation and alterations in airway remodelling.

The epithelium is a dynamic barrier and the damage to this epithelial layer governs a variety of complex mechanisms involving not only epithelial cells but all resident tissue constituents, including immune and stroma cells. Traditionally, diseases characterized by a damaged epithelium have been considered “immunological diseases,” and research efforts aimed at preventing and treating these diseases have primarily focused on immuno-centric therapeutic strategies, that often fail to halt or reverse the natural progression of the disease. In this review, we intend to focus on specific mechanisms driven by the epithelium that ensure barrier function. We will bring asthma and Inflammatory Bowel Diseases into the spotlight, as we believe that these two diseases serve as pertinent examples of epithelium derived pathologies. Finally, we will argue how targeting the epithelium is emerging as a novel therapeutic strategy that holds promise for addressing these chronic diseases.

Ferritin is a cytosolic molecule comprised of subunits that self‐assemble into a nanocage capable of containing up to 4500 iron atoms. Iron stored within ferritin can be mobilized for use within cells or exported from cells. Expression of ferroportin (Fpn) results in export of cytosolic iron and ferritin degradation. Fpn‐mediated iron loss from ferritin occurs in the cytosol and precedes ferritin degradation by the proteasome. Depletion of ferritin iron induces the monoubiquitination of ferritin subunits. Ubiquitination is not required for iron release but is required for disassembly of ferritin nanocages, which is followed by degradation of ferritin by the proteasome. Specific mammalian machinery is not required to extract iron from ferritin. Iron can be removed from ferritin when ferritin is expressed in Saccharomyces cerevisiae , which does not have endogenous ferritin. Expressed ferritin is monoubiquitinated and degraded by the proteasome. Exposure of ubiquitination defective mammalian cells to the iron chelator desferrioxamine leads to degradation of ferritin in the lysosome, which can be prevented by inhibitors of autophagy. Thus, ferritin degradation can occur through two different mechanisms.

Introduction: Asthma is the most common chronic inflammatory disease of the airways. The airway epithelium is a key driver of the disease, and numerous studies have established genome-wide differences in mRNA expression between health and asthma. However, the underlying molecular mechanisms for such differences remain poorly understood. The human TTP family is comprised of ZFP36, ZFP36L1 and ZFP36L2, and has essential roles in immune regulation by determining the stability and translation of myriad mRNAs encoding for inflammatory mediators. We investigated the expression and possible role of the tristetraprolin (TTP) family of RNA binding proteins (RBPs), poorly understood in asthma. Methods: We analysed the levels of ZFP36 , ZFP36L1 and ZFP36L2 mRNA in several publicly available asthma datasets, including single cell RNA-sequencing. We also interrogated the expression of known targets of these RBPs in asthma. We assessed the lung mRNA expression and cellular localization of Zfp36l1 and Zfp36l2 in precision cut lung slices in murine asthma models. Finally, we determined the expression in airway epithelium of ZFP36L1 and ZFP36L2 in human bronchial biopsies and performed rescue experiments in primary bronchial epithelium from patients with severe asthma. Results: We found ZFP36L1 and ZFP36L2 mRNA levels significantly downregulated in the airway epithelium of patients with very severe asthma in different cohorts (5 healthy vs. 8 severe asthma; 36 moderate asthma vs. 37 severe asthma on inhaled steroids vs. 26 severe asthma on oral corticoids). Integrating several datasets allowed us to infer that mRNAs potentially targeted by these RBPs are increased in severe asthma. Zfp36l1 was downregulated in the lung of a mouse model of asthma, and immunostaining of ex vivo lung slices with a dual antibody demonstrated that Zfp36l1/l2 nuclear localization was increased in the airway epithelium of an acute asthma mouse model, which was further enhanced in a chronic model. Immunostaining of human bronchial biopsies showed that airway epithelial cell staining of ZFP36L1 was decreased in severe asthma as compared with mild, while ZFP36L2 was upregulated. Restoring the levels of ZFP36L1 and ZFP36L2 in primary bronchial epithelial cells from patients with severe asthma decreased the mRNA expression of IL6 , IL8 and CSF2 . Discussion: We propose that the dysregulation of ZFP36L1/L2 levels as well as their subcellular mislocalization contributes to changes in mRNA expression and cytoplasmic fate in asthma.

Mon1a has been shown to function in the endolysosomal pathway functioning in the Mon1-Ccz1 complex and it also acts in the secretory pathway where it interacts with dynein and affects ER to Golgi traffic. Here we show that Mon1a is also required for maintenance of the Golgi apparatus. We identified the F-BAR protein FCHO2 as a Mon1a-interacting protein by both yeast two-hybrid analysis and co-immunoprecipitation. siRNA-dependent reductions in Mon1a or FCHO2 resulted in Golgi fragmentation. Membrane trafficking through the secretory apparatus in FCHO2-depleted cells was unaltered, however, reduction of FCHO2 affected the uniform distribution of Golgi enzymes necessary for carbohydrate modification. Fluorescence recovery after photobleaching analysis showed that the Golgi ministacks in Mon1a- or FCHO2-silenced cells did not exchange resident membrane proteins. The effect of FCHO2 silencing on Golgi structure was partially cell cycle-dependent and required mitosis-dependent Golgi fragmentation, whereas the effect of Mon1a-silencing on Golgi disruption was not cell cycle-dependent. mCherry-FCHO2 transiently colocalized on Golgi structures independent of Mon1a. These findings suggest that Mon1a has functions throughout the secretory pathway including interacting with dynein at the ER-Golgi interface in vesicle formation and then interacting with FCHO2 at the Golgi to generate lateral links between ministacks, thus creating Golgi ribbons.Mon1a has been shown to function in the endolysosomal pathway functioning in the Mon1-Ccz1 complex and it also acts in the secretory pathway where it interacts with dynein and affects ER to Golgi traffic. Here we show that Mon1a is also required for maintenance of the Golgi apparatus. We identified the F-BAR protein FCHO2 as a Mon1a-interacting protein by both yeast two-hybrid analysis and co-immunoprecipitation. siRNA-dependent reductions in Mon1a or FCHO2 resulted in Golgi fragmentation. Membrane trafficking through the secretory apparatus in FCHO2-depleted cells was unaltered, however, reduction of FCHO2 affected the uniform distribution of Golgi enzymes necessary for carbohydrate modification. Fluorescence recovery after photobleaching analysis showed that the Golgi ministacks in Mon1a- or FCHO2-silenced cells did not exchange resident membrane proteins. The effect of FCHO2 silencing on Golgi structure was partially cell cycle-dependent and required mitosis-dependent Golgi fragmentation, whereas the effect of Mon1a-silencing on Golgi disruption was not cell cycle-dependent. mCherry-FCHO2 transiently colocalized on Golgi structures independent of Mon1a. These findings suggest that Mon1a has functions throughout the secretory pathway including interacting with dynein at the ER-Golgi interface in vesicle formation and then interacting with FCHO2 at the Golgi to generate lateral links between ministacks, thus creating Golgi ribbons.

The epithelium is a dynamic barrier and the damage to this epithelial layer governs a variety of complex mechanisms involving not only epithelial cells but all resident tissue constituents, including immune and stroma cells. Traditionally, diseases characterized by a damaged epithelium have been considered 'immunological diseases,' and research efforts aimed at preventing and treating these diseases have primarily focused on immuno-centric therapeutic strategies, that often fail to halt or reverse the natural progression of the disease. In this review, we intend to focus on specific mechanisms driven by the epithelium that ensure barrier function. We will bring asthma and Inflammatory Bowel Diseases into the spotlight, as we believe that these two diseases serve as pertinent examples of epithelium derived pathologies. Finally, we will argue how targeting the epithelium is emerging as a novel therapeutic strategy that holds promise for addressing these chronic diseases.

Asthma is deemed an inflammatory disease, yet the defining diagnostic symptom is mechanical bronchoconstriction. We previously discovered a conserved process that drives homeostatic epithelial cell death in response to mechanical cell crowding called cell extrusion(1, 2). Here, we show that the pathological crowding of a bronchoconstrictive attack causes so much epithelial cell extrusion that it damages the airways, resulting in inflammation and mucus secretion. While relaxing airways with the rescue treatment albuterol did not impact these responses, inhibiting live cell extrusion signaling during bronchoconstriction prevented all these symptoms. Our findings propose a new etiology for asthma, dependent on the mechanical crowding of a bronchoconstrictive attack. Our studies suggest that blocking epithelial extrusion, instead of ensuing downstream inflammation, could prevent the feed-forward asthma inflammatory cycle.

Mon1 is an evolutionarily conserved gene that has homologs from yeast to humans. The original identification and characterization of Mon1 in mammals, Mon1a, was performed in a study that identified Mon1a as a modifier of iron homeostasis in mice. That work demonstrated that C57BL mice harbor an intrinsic \"gain-of-function\" mutation that resulted in an excess of the iron exporter ferroportin at the cell surface of iron recycling macrophages. The study also showed that Mon1a had a function in the movement of soluble and membrane-bound proteins through the secretory apparatus. We were able to expand on those findings using protein interaction and RNAi analysis to demonstrate that Mon1a associates with the molecular motor Dynein, known to function in ER-Golgi trafficking. Subcellular localization demonstrated that Mon1a peripherally associates with the ER membrane. Further, RNAi-mediated reduction of Mon1a resulted in a significant decrease in the formation of ER-derived vesicle, which resulted in impaired trafficking in the early secretory pathway. We also determined that the movement of the viral protein VSVGtsGFP from the Golgi to the plasma membrane was delayed in Mon1a-depleted cells. A yeast two-hybrid (Y2H) analysis of Mon1a interacting partners found that a F-BAR domain-contain protein, FCHo2, known to affect membrane traffic at the cell surface, physically associated with Mon1a. RNAi-mediate reduction of Mon1a or FCHo2 resulted in severe Golgi fragmentation, which was dependent on the activity of the Golgi GTPase Rab6. The RNAi-mediate phenotypes of Mon1a and FCHo2 were not identical as only FCHo2 silencing-induced Golgi fragmentation was cell cycle-dependent. We show using FRAP analysis that FCHo2 is necessary for the lateral movement of membrane proteins between Golgi elements that link Golgi cisternae. We determined that FCHo2-mediated Golgi fragmentation resulted in immature glycosylation moieties at the plasma membrane. This dissertation describes novel roles for both Mon1a and FCHo2 in membrane traffic in the secretory pathway and Golgi architecture maintenance.

Epithelial barriers represent the first line of defence against pathogens, yet their role in innate immunity is typ-ically relegated to pathogen detection and immune cell recruitment. This perspective ignores a fundamental evolutionary principle: epithelia defended against pathogens long before complex immune systems evolved. Here, we demonstrate that human bronchial epithelial monolayers retain this ancestral capacity, autono-mously clearing rhinovirus (RV) within 24 hours by selectively extruding infected cells—a process we term virus-induced cell extrusion (VICE). VICE occurs in two waves: a rapid response occurring independently of virus entry, followed by a replication-dependent wave. Barrier-defective epithelia that cannot extrude fail to clear RV. While extrusion maintains barrier integrity and eliminates local infection, it also expels virus-laden cells, promoting transmission. Thus, VICE enables leukocyte-independent epithelial defence while inadver-tently promoting viral transmission, reflecting an evolutionary strategy that prioritizes barrier integrity over containment. These findings redefine epithelia as central players in viral pathogenesis and host protection.

Asthma is a prevalent inflammatory disease marked by life-threatening airway constriction. Current therapies alleviate symptoms by relaxing airway smooth muscle and reducing inflammation but fail to address the underlying airway remodelling, which drives hyper-responsiveness and lung function decline. We previously showed that bronchoconstriction mechanics trigger pathological cell extrusion, wounding the epithelial barrier and perpetuating inflammation. Here, we reveal that a vicious cycle of epithelial damage and inflammation, perpetuates airways in a chronically wounded state. By inhibiting both extrusion and inflammation in mice with established airway asthma symptoms, we synergistically reverse airway remodelling and hyperresponsiveness to methacholine challenge. Moreover, this dual treatment reverts asthmatic transcriptomic and proteomic profiles to healthy states, restoring normal airway function. Our findings offer a novel therapeutic approach to not only halt but reverse asthma progression.