Negar Joharinia, university of Ottawa

Negar Joharinia, Caroline Silveira Falleiros Silva, Vinayahumar Siragam, Mariam Maltseva, Marc-André Langlois. All authors are from Department of Biochemistry, Microbiology & Immunology, Faculty of Medicine, University of Ottawa.

Expressing & Multiplying

Among coronaviruses, the seasonal human coronavirus NL63 (sHCoV-NL63) and SARS-CoV-2 share the cell-surface receptor ACE2 for entry, despite being associated with distinct clinical outcomes, disease severity and degrees of immune dysregulation. Severe COVID-19 is characterized by CD4⁺ T-cell lymphopenia and cellular impairments; however, whether this reflects direct virus–T-cell interactions or secondary inflammatory effects remains incompletely understood. Here, we quantified ACE2 receptor expression on activated primary human CD4⁺ T cells and evaluated expression modulation and downstream immune signaling following NL63 infection. Primary human CD4⁺ T cells were purified by negative selection, activated, and infected with NL63. Activation substantially increased surface ACE2 relative to resting cells. NL63 infection elicited a transient ACE2 upshift at 2 days post-infection (DPI), followed by progressive downregulation at 4 and 7 DPI. ACE2 loss was MOI dependent, with accelerated downmodulation at higher MOI, indicating a dose-responsive effect on receptor regulation. Orthogonal measurements by ddPCR and immunoblotting confirmed concordant reductions in ACE2 mRNA transcripts and protein. In parallel, NL63 triggered early STAT1 and STAT3 phosphorylation at 2-4 DPI, followed by a pronounced reduction of pSTAT1/3 by 7 DPI. Collectively, these data establish activated primary CD4⁺ T cells as cellular targets in which NL63 drives time- and dose-dependent ACE2 downregulation accompanied by biphasic STAT1/3 signaling, highlighting a dynamic interplay between ACE2 modulation and antiviral pathway engagement.