title: A Proteomic-Based Workflow Using Purified Respiratory Syncytial Virus Particles to Identify Cellular Factors as Drug Targets.
abstract: The identification of cellular factors that play a role in respiratory syncytial virus (RSV) replication is an alternative strategy in the identification of druggable cellular protein that are essential for RSV replication. In this regard experimental strategies that are able to screen relevant proteins from the vast array of proteins in the cellular milieu will facilitate the identification of potential drug targets. In this chapter we describe a procedure where RSV particles are purified from cells that are permissive for RSV infection, and the protein composition of the purified virus particles characterized using a proteomics-based strategy. This procedure revealed that actin, several actin-binding proteins, and the chaperones HSP70 and HSP90 also co-purified with the virus particles. The relevance of the HSP90 protein to virus replication was then further validated using imaging, gene silencing and by using an established small molecule HSP90 inhibitor.
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551, Singapore.
authors.forename: Tra Nguyen
authors.initials: TN
authors.lastname: Huong
authors.affiliation: Detection and Diagnostics Laboratory, DSO National Laboratories, 27 Medical Drive, Singapore, 117510, Singapore.
authors.forename: Boon Huan
authors.initials: BH
authors.lastname: Tan
authors.affiliation: School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551, Singapore. rjsugrue@ntu.edu.sg.
authors.forename: Richard J
authors.initials: RJ
authors.lastname: Sugrue
mesh_terms: D000199:Actins / Q000302:isolation & purification; D002460:Cell Line; D002853:Chromatography, Liquid; D018840:HSP70 Heat-Shock Proteins / Q000302:isolation & purification; D018841:HSP90 Heat-Shock Proteins / Q000302:isolation & purification; D006801:Humans; D008840:Microfilament Proteins / Q000302:isolation & purification; D040901:Proteomics / Q000379:methods*; D018357:Respiratory Syncytial Virus Infections / Q000378:metabolism* / Q000821:virology; D018113:Respiratory Syncytial Virus, Human / Q000378:metabolism / Q000502:physiology*; D053719:Tandem Mass Spectrometry; D014771:Virion / Q000378:metabolism / Q000502:physiology*; D014779:Virus Replication
chemical_list: D000199:Actins; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D008840:Microfilament Proteins
keywords: Cell factors; Cytoskeleton; HSP90; LC MS/MS; Paramyxovirus; Proteomics; Respiratory syncytial virus; Virus filaments; Virus morphogenesis