Like many scientists we asked ourselves how we could contribute to stem the surge of SARS-CoV-2 infections that ultimately led to the COVID-19 pandemic. Our lab’s biosafety level is not sufficient to study live human pathogenic viruses. However, we built on the notion that it is not the virus as a whole, but its proteins that hijack host systems and wreak havoc. Early on, several studies were published on the virus-host interactome, providing detailed networks of the SARS-CoV-2 proteins and their human protein interactions. We wondered which of these viral proteins might cause the pathogenicity seen in the varied tissues affected in COVID-19. In our lab, fruit flies (technically vinegar flies, Drosophila melanogaster) are the main model system. The flies are great at providing rapid large-scale screens, be they genetic, molecular or drugs, and provide an unmatched array of genetic tools. In fact, Drosophila has been used successfully in the past to identify pathogenic proteins and the affected pathways for the human HIV-1 and Zika viruses, among others.
First, we set out to identify the SARS-CoV-2 encoded proteins that are most likely pathogenic. The SARS-CoV-2 proteins were prioritized based on toxicity and cellular localization mammalian cells, along with their predicted topology and data from UniProt Knowledgebase. We then generated flies that expressed the most promising proteins ubiquitously. These studies identified four highly pathogenic SARS-CoV-2 proteins; Orf6, Nsp6, Orf3a and Orf7a. The findings and the mechanism underlying Orf6 pathogenicity are described in previous publications (Lee et al. 2021, Cell Biosci.; Zhu et al. 2021, Cell Biosci.). In our latest manuscript we focus on the pathomechanisms of Nsp6, whose expression is highly damaging to the heart. This is of particular interest, as cardiovascular complications are common in patients with COVID-19, and cardiovascular damage has been observed following SARS-CoV-2 infection even in those who remain asymptomatic. In addition, SARS-CoV-2 has been detected in cardiac tissue of patients with COVID-19, suggestion the complications are a direct effect of infection. Notably, the fly has a lengthy and strong track record in modeling heart development and diseases; a major research interest at our Center for Precision Disease Modeling (CPDM). By expressing SARS-CoV-2 Nsp6 specifically in the fly heart we found that Nsp6 induced both structural (disorganized actin filaments and reduced muscle fiber density) and functional (reduced diastolic, but not systolic, diameter of the heart tube; lengthened heart period) defects. We then used proteomics in human cells (HEK 293T) to identify which host proteins can interact with Nsp6, this implicated components of the MGA/MAX complex (MGA, PCGF6 and TFDP1). This complex forms a delicately balanced system with MYC/MAX in regulating the activity of the glycolysis pathway. Complementary transcriptomic data from the fly heart, revealed that Nsp6 binding of the MGA/MAX complex shifts the balance towards MYC/MAX which results in increased glycolysis pathway activity. Additional findings in the flies showed that the Nsp6-induced glycolysis disrupts the function of cardiac mitochondria. Dysfunctional cardiac mitochondria have previously been associated with increased reactive oxygen species (ROS) during heart failure. Altogether, providing a pathomechanism that might explain COVID-19-associated cardiac complications. Furthermore, we found that by inhibiting the glycolysis pathways, using 2-deoxy-D-glucose (2DG) treatment, the Nsp6-induced cardiac pathology in flies could be reduced. To bridge the gap between findings in fly and humans, we validated these findings (pathway affected, phenotype, and 2DG treatment) in mouse primary cardiomyocytes. Currently, several clinical trials using 2DG to treat COVID-19 are ongoing. Moreover, 2DG has received emergency use authorization in India to curb a devastating recent outbreak of COVID-19 (https://pib.gov.in/PressReleasePage.aspx?PRID=1717007).
Altogether these findings point to the glycolysis pathways as a potential target for pharmaceutical intervention in treating cardiac failure associated with COVID-19. In a broader sense, these studies also show the amazing potential of Drosophila in providing a cost-effective, fast, versatile, and highly adaptive systems to study SARS-CoV-2. Making the tiny fly a powerful ally in our fight to protect against coronaviruses.
Authors: Jun-yi Zhu 1,2, Guanglei Wang 3, Xiaohu Huang 1,2, Hangnoh Lee 1,2, Jin-Gu Lee 1,2, Penghua Yang 3, Joyce van de Leemput 1,2, Weiliang Huang 4,5, Maureen A. Kane 4, Peixin Yang 3, and Zhe Han 1,2
1 Center for Precision Disease Modeling, Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA. 2 Division of Endocrinology, Diabetes and Nutrition, Department of Medicine, University of Maryland School of Medicine, Baltimore, MD, USA. 3 Department of Obstetrics, Gynecology & Reproductive Sciences, University of Maryland School of Medicine, Baltimore, MD, USA. 4 Department of Pharmaceutical Sciences, University of Maryland School of Pharmacy, Baltimore, MD, USA. 5 Present address: University of Queensland, Brisbane, QLD 4072, Australia.