Tyrosine sulfation of the amino terminus of PSGL-1 is critical for enterovirus 71 infection

Enterovirus 71 (ev71) is one of the major causative agents of hand,foot,and mouth disease,a common febrile disease in children; however,ev71 has been also associated with various neurological diseases including fatal cases in large ev71 outbreaks particularly in the asia pacific region. recently we identified human p-selectin glycoprotein ligand-1 (psgl-1) as a cellular receptor for entry and replication of ev71 in leukocytes. psgl-1 is a sialomucin expressed on the surface of leukocytes,serves as a high affinity counterreceptor for selectins,and mediates leukocyte rolling on the endothelium. the psgl-1-p-selectin interaction requires sulfation of at least one of three clustered tyrosines and an adjacent o-glycan expressing sialyl lewis x in an n-terminal region of psgl-1. to elucidate the molecular basis of the psgl-1-ev71 interaction,we generated a series of psgl-1 mutants and identified the post-translational modifications that are critical for binding of psgl-1 to ev71. we expressed the psgl-1 mutants in 293t cells and the transfected cells were assayed for their abilities to bind to ev71 by flow cytometry. we found that o-glycosylation on t57,which is critical for psgl-1-selectin interaction,is not necessary for psgl-1 binding to ev71. on the other hand,site-directed mutagenesis at one or more potential tyrosine sulfation sites in the n-terminal region of psgl-1 significantly impaired psgl-1 binding to ev71. furthermore,an inhibitor of sulfation,sodium chlorate,blocked the psgl-1-ev71 interaction and inhibited psgl-1-mediated viral replication of ev71 in jurkat t cells in a dose-dependent manner. thus,the results presented in this study reveal that tyrosine sulfation,but not oglycosylation,in the n-terminal region of psgl-1 may facilitate virus entry and replication of ev71 in leukocytes. © 2010 nishimura et al.