SARS-CoV-2 infection alters mitochondrial and cytoskeletal function in human respiratory epithelial cells mediated by expression of spike protein.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, SCV2), which has resulted in higher morbidity and mortality rate than other respiratory viral infections, such as Influenza A virus (IAV) infection. Investigating the molecular mechanisms of SCV2-host infection vs IAV is vital in exploring antiviral drug targets against SCV2. We assessed differential gene expression in human nasal cells upon SCV2 or IAV infection using RNA sequencing. Compared to IAV, we observed alterations in both metabolic and cytoskeletal pathways suggestive of epithelial remodeling in the SCV2-infected cells, reminiscent of pathways activated as a response to chronic injury. We found that spike protein interaction with the epithelium was sufficient to instigate these epithelial responses using a SCV2 spike pseudovirus. Specifically, we found downregulation of the mitochondrial markers SIRT3 and TOMM22. Moreover, SCV2 spike infection increased extracellular acidification and decreased oxygen consumption rate in the epithelium. In addition, we observed cytoskeletal rearrangements with a reduction in the actin-severing protein cofilin-1 and an increase in polymerized actin, indicating epithelial cytoskeletal rearrangements. This study revealed distinct epithelial responses to SCV2 infection, with early mitochondrial dysfunction in the host cells and evidence of cytoskeletal remodeling that could contribute to the worsened outcome in COVID-19 patients compared to IAV patients. These changes in cell structure and energetics could contribute to cellular resilience early during infection, allowing for prolonged cell survival and potentially paving the way for more chronic symptoms. IMPORTANCE COVID-19 has caused a global pandemic affecting millions of people worldwide, resulting in a higher mortality rate and concerns of more persistent symptoms compared to influenza A. To study this, we compare lung epithelial responses to both viruses. Interestingly, we found that in response to SARS-CoV-2 infection, the cellular energetics changed and there were cell structural rearrangements. These changes in cell structure could lead to prolonged epithelial cell survival, even in the face of not working well, potentially contributing to the development of chronic symptoms. In summary, these findings represent strategies utilized by the cell to survive the infection but result in a fundamental shift in the epithelial phenotype, with potential long-term consequences, which could set the stage for the development of chronic lung disease or long COVID-19.

Pomegranate activates TFEB to promote autophagy-lysosomal fitness and mitophagy.

Mitochondrial dysfunction underscores aging and diseases. Mitophagy (mitochondria + autophagy) is a quality control pathway that preserves mitochondrial health by targeting damaged mitochondria for autophagic degradation. Hence, molecules or compounds that can augment mitophagy are therapeutic candidates to mitigate mitochondrial-related diseases. However, mitochondrial stress remains the most effective inducer of mitophagy. Thus, identification of mitophagy-inducing regimes that are clinically relevant is favorable. In this study, pomegranate extract (PE) supplementation is shown to stimulate mitophagy. PE activates transcription factor EB (TFEB) to upregulate the expression of autophagy and lysosomal genes for mitochondrial quality control under basal and stress conditions. Basally, PE alters mitochondrial morphology and promotes recruitment of autophagosomes to the mitochondria (mitophagosome formation). Upon onset of mitochondrial stress, PE further augments mitophagosome formation, and engages PINK1 and Parkin to the mitochondria to potentiate mitophagy. This cellular phenomenon of PE-induced mitophagy helps to negate superfluous mitochondrial reactive oxygen species (ROS) production and mitochondrial impairment. Overall, our study highlights the potential of PE supplementation as a physiological therapy to modulate TFEB activity to alleviate mitochondrial dysfunction in aging and mitochondrial-related diseases.

Neutrophil TLR4 and PKR are targets of breast cancer cell glycosaminoglycans and effectors of glycosaminoglycan-induced APRIL secretion.

A proliferation-inducing ligand (APRIL), which induces survival and migration signals and tumor growth, is commonly observed in breast cancer tissues but is not often expressed in breast cancer cells themselves. Here, we examined whether breast cancer cells induce APRIL secretion from neutrophils, which are frequently recruited into the breast tumor microenvironment. We found that breast cancer cells do stimulate neutrophils to secrete APRIL through their glycosaminoglycans. Breast cancer cells depleted of heparan sulfate or chondroitin sulfate glycosaminoglycans lose their ability to induce APRIL secretion from neutrophils, and heparan sulfate and chondroitin sulfate can induce secretion that is comparable to that of breast cancer cell-induced secretion. While stimulation of the RNA-activated protein kinase (PKR) is sufficient to induce neutrophil APRIL secretion, both PKR and the toll-like receptor 4 (TLR4) are required for breast cancer cell glycosaminoglycan-induced secretion as separate and specific inhibition of TLR4 or PKR completely prevents the process, suggesting that breast cancer cell glycosaminoglycans target neutrophil TLR4 and PKR to trigger APRIL secretion. Thus, apart from the putative role of cell surface heparan sulfate in binding APRIL that leads to cell growth, we demonstrate that heparan sulfate, as well as chondroitin sulfate plays a novel role in promoting neutrophil secretion of APRIL that could lead to further cell growth. We propose that breast cancer cells take advantage of the neutrophil recruitment to the tumor microenvironment through the dual role of heparan sulfate as cell surface receptor or docking molecule for APRIL and as a ligand that induces neutrophil APRIL secretion to promote their own growth.