Importance of multiple glycan clusters. The dependence of the SARS-CoV-2 antiviral functions of lectins on a set of functionally key glycans in the Spike S1 subunit was demonstrated using pseudotyped viruses containing single glycan attachment site mutants of the Wuhan variant. Mutation of any of the key glycans caused a loss of sensitivity to lectin-based inactivation as judged by an increase in CV-N and GRFT IC 50 values (Table ). The similar but non-identical sets of glycans needed for CV-N and GRFT inhibition of infection likely reflects differences in the engagement of the high-mannose glycans by these two lectins. The individual mutations of these glycan-attachment sites also caused a loss of the irreversible inactivation effect of CV-N (Figure ), suggesting possible cooperative effects between glycans. However, since several of these key glycan-attachment site mutants exhibited greatly reduced infectivity, we cannot dismiss the possibility of more global Spike disruptions, and deciphering this can be addressed by future structural and functional studies.