Structure of mouse coronavirus spike protein complexed with receptor reveals mechanism for viral entry

Abstract

Author summary Coronaviruses recognize many receptors using their envelope-anchored spike protein. The role of receptor binding in coronavirus entry into host cells is a fundamental question in virology. Mouse hepatitis coronavirus (MHV) is unique among all coronaviruses in that it uses the N-terminal domain (NTD) of its spike protein to bind a protein receptor CEACAM1a. While extensive research has been performed on the cell entry mechanisms of coronaviruses that use a different domain of their spike protein for receptor binding, the cell entry mechanism for MHV is still elusive. Here we determined the cryo-EM structure of MHV spike protein complexed with CEACAM1a. The structure reveals unique features of receptor binding by MHV spike that facilitate the structural changes of MHV spike and promote cell entry of MHV. We further confirmed the structural results with biochemical and negative-stain EM analyses. These results suggest that receptor binding plays dual roles in MHV entry: it promotes both viral attachment to host cells and the fusion of host and viral membranes. Our study provides insight into the molecular mechanism of MHV entry, demonstrating how cell entry of MHV has been adapted to its unique way of receptor binding. Coronaviruses recognize a variety of receptors using different domains of their envelope-anchored spike protein. How these diverse receptor recognition patterns affect viral entry is unknown. Mouse hepatitis coronavirus (MHV) is the only known coronavirus that uses the N-terminal domain (NTD) of its spike to recognize a protein receptor, CEACAM1a. Here we determined the cryo-EM structure of MHV spike complexed with mouse CEACAM1a. The trimeric spike contains three receptor-binding S1 heads sitting on top of a trimeric membrane-fusion S2 stalk. Three receptor molecules bind to the sides of the spike trimer, where three NTDs are located. Receptor binding induces structural changes in the spike, weakening the interactions between S1 and S2. Using protease sensitivity and negative-stain EM analyses, we further showed that after protease treatment of the spike, receptor binding facilitated the dissociation of S1 from S2, allowing S2 to transition from pre-fusion to post-fusion conformation. Together these results reveal a new role of receptor binding in MHV entry: in addition to its well-characterized role in viral attachment to host cells, receptor binding also induces the conformational change of the spike and hence the fusion of viral and host membranes. Our study provides new mechanistic insight into coronavirus entry and highlights the diverse entry mechanisms used by different viruses.