Abstract
Phosphatidylinositol kinases (PI kinases) play an important role in the life cycle of several viruses after infection. Using gene knockdown technology, we demonstrate that phosphatidylinositol 4-kinase IIIβ (PI4KB) is required for cellular entry by pseudoviruses bearing the severe acute respiratory syndrome-coronavirus (SARS-CoV) spike protein and that the cell entry mediated by SARS-CoV spike protein is strongly inhibited by knockdown of PI4KB. Consistent with this observation, pharmacological inhibitors of PI4KB blocked entry of SARS pseudovirions. Further research suggested that PI4P plays an essential role in SARS-CoV spike-mediated entry, which is regulated by the PI4P lipid microenvironment. We further demonstrate that PI4KB does not affect virus entry at the SARS-CoV S-ACE2 binding interface or at the stage of virus internalization but rather at or before virus fusion. Taken together, these results indicate a new function for PI4KB and suggest a new drug target for preventing SARS-CoV infection.
Highlights
severe acute respiratory syndrome (SARS)-CoV entry into target cells is a unique and complex process
We demonstrate that phosphatidylinositol 4-kinase III (PI4KB) is required for cellular entry by pseudoviruses bearing the severe acute respiratory syndromecoronavirus (SARS-CoV) spike protein and that the cell entry mediated by SARS-CoV spike protein is strongly inhibited by knockdown of PI4KB
LY294002 and Wortmannin Treatment Inhibit SARS-CoV S-mediated Entry into VeroE6 Cells—To assess the efficiency of virus entry, we produced GFP-containing pseudovirus with SARS-CoV envelope glycoproteins and VSV-G protein (VSVP), with infected cells fluorescing green during microscopy
Summary
SARS-CoV entry into target cells is a unique and complex process. Results: Cell entry mediated by SARS-CoV S was strongly inhibited after knockdown of PI4KB. We demonstrate that phosphatidylinositol 4-kinase III (PI4KB) is required for cellular entry by pseudoviruses bearing the severe acute respiratory syndromecoronavirus (SARS-CoV) spike protein and that the cell entry mediated by SARS-CoV spike protein is strongly inhibited by knockdown of PI4KB. Consistent with this observation, pharmacological inhibitors of PI4KB blocked entry of SARS pseudovirions. We further demonstrate that PI4KB does not affect virus entry at the SARS-CoV S-ACE2 binding interface or at the stage of virus internalization but rather at or before virus fusion Taken together, these results indicate a new function for PI4KB and suggest a new drug target for preventing SARS-CoV infection
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