Dexamethasone sensitizes to ferroptosis by glucocorticoid receptor-induced dipeptidase-1 expression and glutathione depletion.

Dexamethasone is widely used as an immunosuppressive therapy and recently as COVID-19 treatment. Here, we demonstrate that dexamethasone sensitizes to ferroptosis, a form of iron-catalyzed necrosis, previously suggested to contribute to diseases such as acute kidney injury, myocardial infarction, and stroke, all of which are triggered by glutathione (GSH) depletion. GSH levels were significantly decreased by dexamethasone. Mechanistically, we identified that dexamethasone up-regulated the GSH metabolism regulating protein dipeptidase-1 (DPEP1) in a glucocorticoid receptor (GR)-dependent manner. DPEP1 knockdown reversed the phenotype of dexamethasone-induced ferroptosis sensitization. Ferroptosis inhibitors, the DPEP1 inhibitor cilastatin, or genetic DPEP1 inactivation reversed the dexamethasone-induced increase in tubular necrosis in freshly isolated renal tubules. Our data indicate that dexamethasone sensitizes to ferroptosis by a GR-mediated increase in DPEP1 expression and GSH depletion. Together, we identified a previously unknown mechanism of glucocorticoid-mediated sensitization to ferroptosis bearing clinical and therapeutic implications.

Functional landscape of SARS-CoV-2 cellular restriction.

A deficient interferon (IFN) response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection has been implicated as a determinant of severe coronavirus disease 2019 (COVID-19). To identify the molecular effectors that govern IFN control of SARS-CoV-2 infection, we conducted a large-scale gain-of-function analysis that evaluated the impact of human IFN-stimulated genes (ISGs) on viral replication. A limited subset of ISGs were found to control viral infection, including endosomal factors inhibiting viral entry, RNA binding proteins suppressing viral RNA synthesis, and a highly enriched cluster of endoplasmic reticulum (ER)/Golgi-resident ISGs inhibiting viral assembly/egress. These included broad-acting antiviral ISGs and eight ISGs that specifically inhibited SARS-CoV-2 and SARS-CoV-1 replication. Among the broad-acting ISGs was BST2/tetherin, which impeded viral release and is antagonized by SARS-CoV-2 Orf7a protein. Overall, these data illuminate a set of ISGs that underlie innate immune control of SARS-CoV-2/SARS-CoV-1 infection, which will facilitate the understanding of host determinants that impact disease severity and offer potential therapeutic strategies for COVID-19.

Single-cell landscape of immunological responses in patients with COVID-19.

In coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, the relationship between disease severity and the host immune response is not fully understood. Here we performed single-cell RNA sequencing in peripheral blood samples of 5 healthy donors and 13 patients with COVID-19, including moderate, severe and convalescent cases. Through determining the transcriptional profiles of immune cells, coupled with assembled T cell receptor and B cell receptor sequences, we analyzed the functional properties of immune cells. Most cell types in patients with COVID-19 showed a strong interferon-α response and an overall acute inflammatory response. Moreover, intensive expansion of highly cytotoxic effector T cell subsets, such as CD4+ effector-GNLY (granulysin), CD8+ effector-GNLY and NKT CD160, was associated with convalescence in moderate patients. In severe patients, the immune landscape featured a deranged interferon response, profound immune exhaustion with skewed T cell receptor repertoire and broad T cell expansion. These findings illustrate the dynamic nature of immune responses during disease progression.

Network perturbation analysis in human bronchial epithelial cells following SARS-CoV2 infection.

BACKGROUND: SARS-CoV2, the agent responsible for the current pandemic, is also causing respiratory distress syndrome (RDS), hyperinflammation and high mortality. It is critical to dissect the pathogenetic mechanisms in order to reach a targeted therapeutic approach. METHODS: In the present investigation, we evaluated the effects of SARS-CoV2 on human bronchial epithelial cells (HBEC). We used RNA-seq datasets available online for identifying SARS-CoV2 potential genes target on human bronchial epithelial cells. RNA expression levels and potential cellular gene pathways have been analyzed. In order to identify possible common strategies among the main pandemic viruses, such as SARS-CoV2, SARS-CoV1, MERS-CoV, and H1N1, we carried out a hypergeometric test of the main genes transcribed in the cells of the respiratory tract exposed to these viruses. RESULTS: The analysis showed that two mechanisms are highly regulated in HBEC: the innate immunity recruitment and the disassembly of cilia and cytoskeletal structure. The granulocyte colony-stimulating factor (CSF3) and dynein heavy chain 7, axonemal (DNAH7) represented respectively the most upregulated and downregulated genes belonging to the two mechanisms highlighted above. Furthermore, the carcinoembryonic antigen-related cell adhesion molecule 7 (CEACAM7) that codifies for a surface protein is highly specific of SARS-CoV2 and not for SARS-CoV1, MERS-CoV, and H1N1, suggesting a potential role in viral entry. In order to identify potential new drugs, using a machine learning approach, we highlighted Flunisolide, Thalidomide, Lenalidomide, Desoximetasone, xylazine, and salmeterol as potential drugs against SARS-CoV2 infection. CONCLUSIONS: Overall, lung involvement and RDS could be generated by the activation and down regulation of diverse gene pathway involving respiratory cilia and muscle contraction, apoptotic phenomena, matrix destructuration, collagen deposition, neutrophil and macrophages recruitment.

LY6E impairs coronavirus fusion and confers immune control of viral disease.

Zoonotic coronaviruses (CoVs) are substantial threats to global health, as exemplified by the emergence of two severe acute respiratory syndrome CoVs (SARS-CoV and SARS-CoV-2) and Middle East respiratory syndrome CoV (MERS-CoV) within two decades1-3. Host immune responses to CoVs are complex and regulated in part through antiviral interferons. However, interferon-stimulated gene products that inhibit CoVs are not well characterized4. Here, we show that lymphocyte antigen 6 complex, locus E (LY6E) potently restricts infection by multiple CoVs, including SARS-CoV, SARS-CoV-2 and MERS-CoV. Mechanistic studies revealed that LY6E inhibits CoV entry into cells by interfering with spike protein-mediated membrane fusion. Importantly, mice lacking Ly6e in immune cells were highly susceptible to a murine CoV-mouse hepatitis virus. Exacerbated viral pathogenesis in Ly6e knockout mice was accompanied by loss of hepatic immune cells, higher splenic viral burden and reduction in global antiviral gene pathways. Accordingly, we found that constitutive Ly6e directly protects primary B cells from murine CoV infection. Our results show that LY6E is a critical antiviral immune effector that controls CoV infection and pathogenesis. These findings advance our understanding of immune-mediated control of CoV in vitro and in vivo-knowledge that could help inform strategies to combat infection by emerging CoVs.

Human Gene Sequences in SARS-CoV-2 and Other Viruses.

In a previous study, we identified a 117 base severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) sequence in the human genome with 94.6% identity. The sequence was in chromosome 1p within an intronic region of the netrin G1 (NTNG1) gene. The sequence matched a sequence in the SARS-CoV-2 Orf1b gene in non-structural protein 14 (NSP14), which is an exonuclease and NSP15, an endoribonuclease. In the current study we compared the human genome with other viral genomes to determine some of the characteristics of human sequences found in the latter. Most of the viruses had human sequences, but they were short. Hepatitis A and St Louis encephalitis had human sequences that were longer than the 117 base SARS-Cov-2 sequence, but they were in non-coding regions of the human genome. The SARS-Cov-2 sequence was the only long sequence found in a human gene (NTNG1). The related coronaviruses SARS-Cov had a 41 BP human sequence on chromosome 3 that was not part of a human gene, and MERS had no human sequence. The 117 base SARS-CoV-2 human sequence is relatively close to the viral spike sequence, separated only by NSP16, a 904 base sequence. The mechanism for SARS-CoV-2 infection is the binding of the virus spike protein to the membrane-bound form of angiotensin-converting enzyme 2 (ACE2) and internalization of the complex by the host cell. We have no explanation for the NSP14 and NSP15 SARS-Cov-2 sequences we observed here or how they might relate to infectiousness. Further studies are warranted.