HCV and SARS-CoV-2 Envelope Glycoproteins Down-Modulate NF-κB Activity in Association with Induction of ER Stress

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University of Liverpool
Viruses, as highly adaptable pathogens, possess the remarkable capability to modulate host immune responses and influence disease pathogenesis. Within this context, our research endeavors focused on studying the intricate interplay between HCV E1E2 Env glycoprotein on downmodulating HIV-1 long terminal repeat (LTR) activation, as well as host gene expression patterns. Measuring the HIV-1 LTR activity in hepatic cells co-transfected with HCV E1E2 Env protein and HIV-1 promoter driving expression of a Luciferase reporter molecule demonstrated that the protein downmodulated LTR activity. Using different reporter constructs we demonstrated that the mechanism is mediated specifically via NF-κB. Notably, within this investigation, it is important to highlight that the use of the HIV-1 LTR as a tool is specifically designed for monitoring NF-κB activation and does not imply co-infection of HCV and HIV cells in vivo. Cells expressing the HCV E1E2 Env glycoprotein, NF-κB relinquished its binding affinity to the HIV-1 LTR promoter, while other transcription factors exhibited augmented interactions. Furthermore, transcriptomic analysis of cells expressing the HCV E1E2 Env glycoprotein identified the stimulation of the endoplasmic reticulum (ER) stress response pathway, concomitant with the upregulation of responsive genes, including ATF3. Through shRNA mediated inhibition of ATF3 expression, we identified that the E1E2 mediated inhibitory effects on HIV-1 LTR activity was alleviated. Additionally, we observe that ATF3 knock-down raises the basal level of HIV-1 LTR activity in the absence of E1E2 Env expression. The result suggested that the HCV Env glycoprotein inhibits NF-κB signalling via upregulation of the ER stress pathway. In parallel to our investigation on HCV E1E2 Env, we extended our molecular approach to explore the impact of SARS-CoV-2 structural proteins, spike (S), membrane (M), envelope (E), and nucleocapsid (N), on HIV-1 LTR activity. We found that Env glycoproteins of SARS-CoV-2 downmodulate LTR activity, while the N protein did not exhibit any discernible impact on LTR activity. Comparative transcriptomic analysis of stable cell lines expressing individual SARS-CoV-2 structural proteins revealed that the viral proteins S, M, and E have the capacity to influence key transcriptional factors, including NF-κB and SP1. Additionally, the expression of these Env glycoproteins stimulated ER stress and activation of the unfolded protein response (UPR). Furthermore, there was an upregulation of HSF-1, a transcription factor involved in regulating heat shock proteins (HSPs) and early gene response. The activation of HSF-1 may play a significant role in counteracting acute inflammation by inhibiting NF-κB and downstream pro-inflammatory signals. Importantly, this response was absent in the context of the N protein, possibly indicating that the N protein may not stimulate HSPs to the same extent as the other SARS-CoV-2 structural proteins, possibly insufficient to activate HSF-1. Together, these findings suggest a potential link between the two viral systems, highlighting common mechanisms by which viral proteins modulate host cellular processes. Both the HCV E1E2 and SARS-CoV-2 Env glycoproteins can perturb NF-κB signalling, leading to alterations in gene expression patterns. This indicates that these viral proteins may exploit similar strategies to evade host immune responses and promote viral replication. This study adds to the growing body of knowledge by providing novel insights into the dynamic interaction between HCV and SARS-CoV-2 viral systems and their impact on host cellular processes. By revealing shared mechanisms of viral protein modulation and their implications for viral pathogenesis and immune responses, this work presents a novel contribution to our understanding of viral infections and paves the way for future investigations and targeted interventions.
HCV E1E2 Env glycoprotein, HIV-1 long terminal repeat (LTR) activation, Luciferase reporter molecule, NF-κB, Transcriptomic analysis, Endoplasmic reticulum (ER) stress response pathway, ATF3, shRNA, SARS-CoV-2 structural proteins, Spike (S), Membrane (M), Envelope (E), Nucleocapsid (N), Comparative transcriptomic analysis, SP1, Unfolded protein response (UPR), HSF-1, Heat shock proteins (HSPs), Viral protein modulation, Host cellular processes, Immune responses, Viral pathogenesis, Acute inflammation.
HCV E1E2, HIV-1 LTR, NF-κB, Transcriptomics, ER stress, ATF3, SARS-CoV-2 proteins, Envelope