ABSTRACT
SARS-CoV-2 spike requires proteolytic processing for viral entry. The presence of a polybasic furin-cleavage site (FCS) in spike, and evolution towards an optimised FCS by dominant variants of concern (VOCs), are linked to enhanced infectivity and transmission. Guanylate binding proteins (GBP) are interferon-inducible restriction factors that target furin-mediated processing of viral envelope proteins and limit infectivity. Here we investigated whether GBPs restrict SARS-CoV-2 infection, and whether VOCs have evolved spikes that escape restriction. We show that GBP2 and 5 interfere with cleavage of the spike proteins of Wuhan-Hu-1, Alpha, Delta and Omicron, consistent with furin inhibition by GBPs. However, while GBP2/5 restrict Wuhan-Hu-1 infectivity, Alpha and Delta escape restriction. GBP exposure in producer cells influences viral entry route into target cells, with a shift towards endosomal entry. We therefore investigated whether GBP-targeting of spike alters sensitivity to endosomal restriction factors, IFITMs. We find IFITM1, but not IFITM 2 or 3, inhibit infection of naturally-permissive epithelial cells by early-lineage SARS-CoV-2, as well as Alpha and Delta, however GBPs did not sensitise to IFITM restriction. Strikingly, we find Omicron is unique amongst VOCs, being sensitive to restriction by GBP2/5, and also IFITM1, 2 and 3. We conclude evolution of Alpha and Delta spikes have conferred resistance to GBP restriction, but this is not solely due to acquisition of an enhanced FCS. Notably, Omicron, which has evolved under different selective pressures, has selected for changes in spike that not only mediate antibody escape, and shift in cell tropism and entry, but also impact the sensitivity of Omicron to innate immunity, potentially contributing to altered pathogenesis.