SARS-CoV-2 targets NKG2D ligands to evade natural killer cell-mediated immunity

Abstract

Severe acute respiratory syndrome (SARS-CoV-2) is responsible for the ongoing global COVID-19 pandemic. This virus continues to drive research to develop vaccines and therapeutics as well as to characterize host immune responses and elucidate mechanisms of viral immune evasion. Natural killer (NK) cells are innate cytotoxic lymphocytes that serve as a first-line defense against viruses. NK cells express a highly potent activating receptor, NKG2D, that is highly conserved and binds to several ligands, namely, MIC-A, MIC-B, and ULBP1-6. These ligands are expressed in response to cellular stress such as viral infection and malignant transformation to trigger NKG2D-mediated NK-cell killing. Many viruses and cancers are known to downmodulate NKG2D ligands to escape immune recognition and viral clearance from their hosts, but studies addressing mechanisms or genes specific to SARS-CoV-2 are lacking. In this work, we found that SARS-CoV-2 infection of the human lung epithelial cell line, A549-ACE2, downregulates NKG2D ligands via shedding from the cell surface. We observed that matrix metalloproteases, which function as sheddases, and MICB are transcriptionally upregulated in infected cells. Upon assessing the clinical relevance of these findings, we observed increased serum MIC-A and MIC-B in COVID-19 patients compared to healthy donors, with serum MIC-A associating with COVID-19 severity. A monoclonal antibody, 7C6, which has been shown to block MIC-A/B cleavage and shedding in pre-clinical cancer studies, increased MIC-A/B surface levels in SARS-CoV-2-infected A549-ACE2 cells, while simultaneously decreasing supernatant levels of MIC-A in a dose-dependent manner. Moreover, 7C6 augmented NK-cell killing and degranulation toward infected cells, highlighting its therapeutic potential. Given the substantial polymorphism of MICA among humans, we assessed the ability of SARS-CoV-2 to downregulate different MIC-A alleles and demonstrated that, while SARS-CoV-2 selectively downregulates surface expression of human MIC-A*001, *002, and 004, MIC-A008 is highly resistant to this effect. Comprehensive screening of all SARS-CoV-2 proteins revealed that only two proteins, Nsp1 and ORF6, were sufficient to downregulate MIC-A/B in A549-ACE2 cells. However, the allele-specific downregulation of MIC-A seen with live virus infection could only be recapitulated with ORF6, suggesting that this is the viral immunevasin responsible for SARS-CoV-2-mediated MIC-A/B downregulation and shedding. Altogether, we find that SARS-CoV-2 downregulates MIC-A/B by inducing shedding from the surface of infected cells. This mechanism of immune evasion can be blocked by 7C6. We propose the use of 7C6 in vivo during SARS-CoV-2 infections to therapeutically enhance NK-cell surveillance and boost viral clearance. We also highlight the need for further study into MICA genotype associations with COVID-19 susceptibility and disease severity.