title: Recombinant respiratory syncytial virus F protein expression is hindered by inefficient nuclear export and mRNA processing.
abstract: Studies of the fusion activity of respiratory syncytial virus (RSV) F protein are significantly hindered by low recombinant expression levels. While infection produces F protein levels detectable by western blot, recombinant expression produces undetectable to low levels of F protein. Identifying the obstacles that hinder recombinant F protein expression may lead to improved expression and facilitate the study of F protein function. We hypothesized that nuclear localization and/or inefficient RNA polymerase II-mediated transcription contribute to poor recombinant F protein expression. This study shows a combination of stalled nuclear export, premature polyadenylation, and low mRNA abundance all contribute to low recombinant F protein expression levels. In addition, this study provides an expression optimization strategy that results in greater F protein expression levels than observed by codon-optimization of the F protein gene, which will be useful for future studies of F protein function.
authors.affiliation: MedImmune, One MedImmune Way, Gaithersburg, MD 20878, USA. huangk@medimmune.com
authors.forename: Kelly
authors.initials: K
authors.lastname: Huang
authors.forename: Heather
authors.initials: H
authors.lastname: Lawlor
authors.forename: Roderick
authors.initials: R
authors.lastname: Tang
authors.forename: Randall S
authors.initials: RS
authors.lastname: MacGill
authors.forename: Nancy D
authors.initials: ND
authors.lastname: Ulbrandt
authors.forename: Herren
authors.initials: H
authors.lastname: Wu
mesh_terms: D021581:Active Transport, Cell Nucleus; D000818:Animals; D002460:Cell Line; D015870:Gene Expression; D006801:Humans; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014760:Viral Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: C526850:F protein, human respiratory syncytial virus; D012333:RNA, Messenger; D012367:RNA, Viral; D011994:Recombinant Proteins; D014760:Viral Fusion Proteins