BMB-11 Establishment of a Respiratory Model for the Study of SARS-CoV-1 Nsp1 Intracellular Functions
Abstract
The COVID-19 pandemic intensely highlighted the devastating impact of coronavirus-family infections, with two notable coronaviruses, Severe Acute Respiratory Syndrome Coronavirus 1 and 2 (SARS-CoV-1 and SARS-CoV-2) capable of causing significant human disease. One potential underlying cause of coronavirus severe pathology is the production of virus-encoded proteins that hijack host cell machinery, allowing for extensive viral proliferation and evasion of the host immune response. One such protein, SARS-CoV-1 non-structural protein 1 (Nsp1), maintains two defined functions during viral replication: (1) inhibition of host protein translation and (2) dysregulation of the Type I interferon antiviral response, a key step to evading the host’s immune response. A potential mechanism by which Nsp1 could achieve these functions is through interaction with host regulators such as ADAR1, which localizes to stress granules during infection. To study the intracellular functions of Nsp1, eukaryotic expression plasmids encoding wild-type SARS-CoV-1 Nsp1 and three functional Nsp1 mutants were generated: M4, which lacks both translational and immune inhibition functions; M16, which lacks immune inhibition alone; and M27, which has increased translational and immune inhibition. Wild-type and mutant Nsp1 were transfected into the human lung epithelial cell line, A549, to model a respiratory infection. Using fluorescent microscopy, Nsp1 expression in the respiratory cells was confirmed along with expression of a GFP-tagged ADAR1, and stress granule formation. Through application and study within this model, we can gain an understanding of the mechanisms by which SARS-CoV-1 evades immune responses. Lessons learned from this work can be employed in the development of novel therapeutics against coronavirus infections and improve patient outcomes during current or future coronavirus outbreaks.
BMB-11 Establishment of a Respiratory Model for the Study of SARS-CoV-1 Nsp1 Intracellular Functions
University Readiness Center Greatroom
The COVID-19 pandemic intensely highlighted the devastating impact of coronavirus-family infections, with two notable coronaviruses, Severe Acute Respiratory Syndrome Coronavirus 1 and 2 (SARS-CoV-1 and SARS-CoV-2) capable of causing significant human disease. One potential underlying cause of coronavirus severe pathology is the production of virus-encoded proteins that hijack host cell machinery, allowing for extensive viral proliferation and evasion of the host immune response. One such protein, SARS-CoV-1 non-structural protein 1 (Nsp1), maintains two defined functions during viral replication: (1) inhibition of host protein translation and (2) dysregulation of the Type I interferon antiviral response, a key step to evading the host’s immune response. A potential mechanism by which Nsp1 could achieve these functions is through interaction with host regulators such as ADAR1, which localizes to stress granules during infection. To study the intracellular functions of Nsp1, eukaryotic expression plasmids encoding wild-type SARS-CoV-1 Nsp1 and three functional Nsp1 mutants were generated: M4, which lacks both translational and immune inhibition functions; M16, which lacks immune inhibition alone; and M27, which has increased translational and immune inhibition. Wild-type and mutant Nsp1 were transfected into the human lung epithelial cell line, A549, to model a respiratory infection. Using fluorescent microscopy, Nsp1 expression in the respiratory cells was confirmed along with expression of a GFP-tagged ADAR1, and stress granule formation. Through application and study within this model, we can gain an understanding of the mechanisms by which SARS-CoV-1 evades immune responses. Lessons learned from this work can be employed in the development of novel therapeutics against coronavirus infections and improve patient outcomes during current or future coronavirus outbreaks.