Abstract SARS-CoV-2 can infect various human tissues and cell types, principally via interaction with its cognate receptor ACE2. However, how the virus evolves in different cellular environments is poorly understood. Here, we used experimental evolution to study the adaptation of the SARS-CoV-2 spike to four human cell lines expressing different levels of key entry factors. After 20 passages, cell type-specific phenotypic changes were observed. Selected spike mutations were identified and functionally characterized in terms of entry efficiency, ACE2 affinity, spike processing, TMPRSS2 usage, entry pathway and syncytia formation. We found that the effects of these mutations varied across cell types. Interestingly, two spike mutations (L48S and A372T) that emerged in cells expressing low ACE2 levels increased receptor affinity, syncytia induction, and entry efficiency under low-ACE2 conditions. Our results demonstrate specific adaptation of the SARS-CoV-2 spike to different cell types and have implications for understanding SARS-CoV-2 tissue tropism and evolution.

Cell type-specific adaptation of the SARS-CoV-2 spike