Identification of novel EV71-interacting proteins by glycoproteomic approaches

  • 蘇 珮誼

Student thesis: Doctoral Thesis


Enterovirus 71 (EV71) is a major causative agent of hand-foot-and-mouth disease (HFMD) EV71 infection of the central nervous system (CNS) causes severe neurological complications in infants Although two highly glycosylated membrane proteins SCARB2 and PSGL-1 have been identified as the cellular and functional receptors of EV71 the pathogenesis of EV71 remains mostly ambiguous The influence of glycosylation on viral infections has been discussed in relation to other picornaviruses Based on these previous findings we first attempted to establish whether cell surface sialylation can mediate EV71-protein recognition and to determine whether the interaction between SCARB2 and EV71 is retarded after desialylation Second the multiple receptor characteristics of EV71 have been explored and it has been suggested that there are unknown EV71-interacting proteins on the cell surface which are involved in the attachment and infection of EV71 We therefore applied glycoproteomic approaches to identify novel EV71-interacting proteins and to verify the roles of candidate proteins in EV71 binding and infection We isolated the EV71-associated glycoproteins for the purpose of protein identification Among the sixteen proteins identified using LC MS/MS spectrometry analysis we selected cell-surface nucleolin for in-depth analysis We found that EV71 interacts directly with nucleolin via the VP1 capsid protein In addition it was found that the knockdown of cell surface nucleolin decreased EV71 binding infection as well as viral infectivity in human cells In addition over-expression of human nucleolin on the surface of murine cells increased EV71 attachment and entry Furthermore we found that the surface nucleolin lactoferrin and plasminogen may associate with SCARB2 and assist EV71 infection Our data strongly suggest that human nucleolin is an receptor for EV71 infection Our work not only contributes to an understanding of virus-receptor interactions in the early stages of EV71 infection but also demonstrates that glycoproteomic technology is a reliable methodology by which to discover novel pathogen-associated proteins
Date of Award2015 Mar 26
Original languageEnglish
SupervisorChuan-Fa Chang (Supervisor)

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