RNA Sequencing of VeroE6 cells infected with SARS-CoV-2 in presence and absence of GYY4137

Viruses modulate various aspects of host physiology, including carbon metabolism, redox balance, and mitochondrial bioenergetics, to acquire the building blocks for replication and regulate the immune response. Understanding newer ways of how SARS-CoV-2 alters host redox metabolism, carbon metabolism, and mitochondrial health may lead to potential treatments for COVID-19. Reports have shown direct corelation of H2S metabolism with SARS-CoV-2 disease severity. However, the molecular mechanism of how H2S controls virus replication remains unknown. In this study, we have attempted to delineate the pathways altered by SARS-CoV-2 infection, using multi-omics techniques, and effect of H2S donor GYY4137 in the same. We have infected VeroE6 cells with SARS-CoV-2 (HK variant) at an MOI of 0.01 for 48 hours in presence and absence of 5 mM GYY4137. The transcriptomic profiles of uninfected, infected and treated were compared, to understand the molecular pathways underlying and to understand how H2S can modulate host metabolism to control viral replication. The study involves RNA sequencing from RNA isolated from VeroE6 cells infected with SARS-CoV-2 in presence or absence of potent drug molecule GYY4137, a slow releasing H2S donor at 48 hours post-infection. The profiles of Infected samples were compared with untreated, while those of treated were compared with untreated(but infected) samples. Samples for all three conditions were prepared in triplicates. Upon SARS-CoV-2 infection, the major pathways upregulated were for cellular response towards viral infection. Signalling pathways like RIG-1 pathway, MAPK pathway, TLR signalling, Cytokine-chemokine signalling were highly enriched with infection. Viral infections have been shown to hijack host metabolism. In line with this, we have observed, deregulation of host central carbon metabolism pathways like glycolysis, PPP, OXPHOS, TCA cycle. We noticed that the treatment of GYY4137 moderately reversed the effect of SARS-CoV-2 infection on the expression of genes associated with the oxidoreductase complex and Nrf2- antioxidant pathway. Furthermore, pathways related to defence and cellular response to viruses, including TLR, TNF, Nod-like receptor, MAPK, RIG-1, and cytokine/chemokine signaling, were more induced in GYY4137 treated VeroE6 cells as compared to untreated cells. We did not observe any effect of GYY4137 on the expression of central metabolism, respiration, and amino acid metabolism. We validated the expression changes of several Nrf2-specific antioxidant genes in response to GYY4137 by qRT-PCR. The qRT-PCR and RNA-seq results were highly consistent and indicated that antioxidant genes were downregulated by SARS-CoV-2 infection, whereas GYY4137 treatment reversed this effect. Altogether, H2S supplementation induces a major realignment of redox metabolism and immune pathways associated with SARS-CoV-2 infection.