SARS-CoV-2 Nsp6 damages Drosophila heart and mouse cardiomyocytes through MGA/MAX complex-mediated increased glycolysis.

SARS-CoV-2 infection causes COVID-19, a severe acute respiratory disease associated with cardiovascular complications including long-term outcomes. The presence of virus in cardiac tissue of patients with COVID-19 suggests this is a direct, rather than secondary, effect of infection. Here, by expressing individual SARS-CoV-2 proteins in the Drosophila heart, we demonstrate interaction of virus Nsp6 with host proteins of the MGA/MAX complex (MGA, PCGF6 and TFDP1). Complementing transcriptomic data from the fly heart reveal that this interaction blocks the antagonistic MGA/MAX complex, which shifts the balance towards MYC/MAX and activates glycolysis-with similar findings in mouse cardiomyocytes. Further, the Nsp6-induced glycolysis disrupts cardiac mitochondrial function, known to increase reactive oxygen species (ROS) in heart failure; this could explain COVID-19-associated cardiac pathology. Inhibiting the glycolysis pathway by 2-deoxy-D-glucose (2DG) treatment attenuates the Nsp6-induced cardiac phenotype in flies and mice. These findings point to glycolysis as a potential pharmacological target for treating COVID-19-associated heart failure.

Drosophila, a powerful model to study virus-host interactions and pathogenicity in the fight against SARS-CoV-2.

The COVID-19 pandemic is having a tremendous impact on humanity. Although COVID-19 vaccines are showing promising results, they are not 100% effective and resistant mutant SARS-CoV-2 strains are on the rise. To successfully fight against SARS-CoV-2 and prepare for future coronavirus outbreaks, it is essential to understand SARS-CoV-2 protein functions, their host interactions, and how these processes convey pathogenicity at host tissue, organ and systemic levels. In vitro models are valuable but lack the physiological context of a whole organism. Current animal models for SARS-CoV-2 research are exclusively mammals, with the intrinsic limitations of long reproduction times, few progeny, ethical concerns and high maintenance costs. These limitations make them unsuitable for rapid functional investigations of virus proteins as well as genetic and pharmacological screens. Remarkably, 90% of the SARS-CoV-2 virus-host interacting proteins are conserved between Drosophila and humans. As a well-established model system for studying human diseases, the fruit fly offers a highly complementary alternative to current mammalian models for SARS-CoV-2 research, from investigating virus protein function to developing targeted drugs. Herein, we review Drosophila's track record in studying human viruses and discuss the advantages and limitations of using fruit flies for SARS-CoV-2 research. We also review studies that already used Drosophila to investigate SARS-CoV-2 protein pathogenicity and their damaging effects in COVID-19 relevant tissues, as well as studies in which the fly was used as an efficient whole animal drug testing platform for targeted therapeutics against SARS-CoV-2 proteins or their host interacting pathways.

Understanding Individual SARS-CoV-2 Proteins for Targeted Drug Development against COVID-19.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of the COVID-19 pandemic, responsible for millions of deaths globally. Even with effective vaccines, SARS-CoV-2 will likely maintain a hold in the human population through gaps in efficacy, percent vaccinated, and arising new strains. Therefore, understanding how SARS-CoV-2 causes widespread tissue damage and the development of targeted pharmacological treatments will be critical in fighting this virus and preparing for future outbreaks. Herein, we summarize the progress made thus far by using in vitro or in vivo models to investigate individual SARS-CoV-2 proteins and their pathogenic mechanisms. We have grouped the SARS-CoV-2 proteins into three categories: host entry, self-acting, and host interacting. This review focuses on the self-acting and host-interacting SARS-CoV-2 proteins and summarizes current knowledge on how these proteins promote virus replication and disrupt host systems, as well as drugs that target the virus and virus interacting host proteins. Encouragingly, many of these drugs are currently in clinical trials for the treatment of COVID-19. Future coronavirus outbreaks will most likely be caused by new virus strains that evade vaccine protection through mutations in entry proteins. Therefore, study of individual self-acting and host-interacting SARS-CoV-2 proteins for targeted therapeutic interventions is not only essential for fighting COVID-19 but also valuable against future coronavirus outbreaks.

Characterization of SARS-CoV-2 proteins reveals Orf6 pathogenicity, subcellular localization, host interactions and attenuation by Selinexor.

BACKGROUND: SARS-CoV-2 causes COVID-19 which has a widely diverse disease profile. The mechanisms underlying its pathogenicity remain unclear. We set out to identify the SARS-CoV-2 pathogenic proteins that through host interactions cause the cellular damages underlying COVID-19 symptomatology. METHODS: We examined each of the individual SARS-CoV-2 proteins for their cytotoxicity in HEK 293 T cells and their subcellular localization in COS-7 cells. We also used Mass-Spec Affinity purification to identify the host proteins interacting with SARS-CoV-2 Orf6 protein and tested a drug that could inhibit a specific Orf6 and host protein interaction. RESULTS: We found that Orf6, Nsp6 and Orf7a induced the highest toxicity when over-expressed in human 293 T cells. All three proteins showed membrane localization in COS-7 cells. We focused on Orf6, which was most cytotoxic and localized to the endoplasmic reticulum, autophagosome and lysosomal membranes. Proteomics revealed Orf6 interacts with nucleopore proteins (RAE1, XPO1, RANBP2 and nucleoporins). Treatment with Selinexor, an FDA-approved inhibitor for XPO1, attenuated Orf6-induced cellular toxicity in human 293 T cells. CONCLUSIONS: Our study revealed Orf6 as a highly pathogenic protein from the SARS-CoV-2 genome, identified its key host interacting proteins, and Selinexor as a drug candidate for directly targeting Orf6 host protein interaction that leads to cytotoxicity.

Functional analysis of SARS-CoV-2 proteins in Drosophila identifies Orf6-induced pathogenic effects with Selinexor as an effective treatment.

BACKGROUND: SARS-CoV-2 causes COVID-19 with a widely diverse disease profile that affects many different tissues. The mechanisms underlying its pathogenicity in host organisms remain unclear. Animal models for studying the pathogenicity of SARS-CoV-2 proteins are lacking. METHODS: Using bioinformatic analysis, we found that 90% of the virus-host interactions involve human proteins conserved in Drosophila. Therefore, we generated a series of transgenic fly lines for individual SARS-CoV-2 genes, and used the Gal4-UAS system to express these viral genes in Drosophila to study their pathogenicity. RESULTS: We found that the ubiquitous expression of Orf6, Nsp6 or Orf7a in Drosophila led to reduced viability and tissue defects, including reduced trachea branching as well as muscle deficits resulting in a "held-up" wing phenotype and poor climbing ability. Furthermore, muscles in these flies showed dramatically reduced mitochondria. Since Orf6 was found to interact with nucleopore proteins XPO1, we tested Selinexor, a drug that inhibits XPO1, and found that it could attenuate the Orf6-induced lethality and tissue-specific phenotypes observed in flies. CONCLUSIONS: Our study established Drosophila as a model for studying the function of SARS-CoV2 genes, identified Orf6 as a highly pathogenic protein in various tissues, and demonstrated the potential of Selinexor for inhibiting Orf6 toxicity using an in vivo animal model system.