title: Recruitment of RED-SMU1 complex by Influenza A Virus RNA polymerase to control Viral mRNA splicing.
abstract: Influenza A viruses are major pathogens in humans and in animals, whose genome consists of eight single-stranded RNA segments of negative polarity. Viral mRNAs are synthesized by the viral RNA-dependent RNA polymerase in the nucleus of infected cells, in close association with the cellular transcriptional machinery. Two proteins essential for viral multiplication, the exportin NS2/NEP and the ion channel protein M2, are produced by splicing of the NS1 and M1 mRNAs, respectively. Here we identify two human spliceosomal factors, RED and SMU1, that control the expression of NS2/NEP and are required for efficient viral multiplication. We provide several lines of evidence that in infected cells, the hetero-trimeric viral polymerase recruits a complex formed by RED and SMU1 through interaction with its PB2 and PB1 subunits. We demonstrate that the splicing of the NS1 viral mRNA is specifically affected in cells depleted of RED or SMU1, leading to a decreased production of the spliced mRNA species NS2, and to a reduced NS2/NS1 protein ratio. In agreement with the exportin function of NS2, these defects impair the transport of newly synthesized viral ribonucleoproteins from the nucleus to the cytoplasm, and strongly reduce the production of infectious influenza virions. Overall, our results unravel a new mechanism of viral subversion of the cellular splicing machinery, by establishing that the human splicing factors RED and SMU1 act jointly as key regulators of influenza virus gene expression. In addition, our data point to a central role of the viral RNA polymerase in coupling transcription and alternative splicing of the viral mRNAs.
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Fournier
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Chiayn
authors.initials: C
authors.lastname: Chiang
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Sandie
authors.initials: S
authors.lastname: Munier
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Andru
authors.initials: A
authors.lastname: Tomoiu
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Caroline
authors.initials: C
authors.lastname: Demeret
authors.affiliation: CNRS, UMR 3569, Paris, France; Institut Pasteur, Unité de Génomique Virale et Vaccination, Département de Virologie, Paris, France.
authors.forename: Pierre-Olivier
authors.initials: PO
authors.lastname: Vidalain
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Yves
authors.initials: Y
authors.lastname: Jacob
authors.affiliation: Institut Pasteur, Unité de Génétique Moléculaire des Virus à ARN, Département de Virologie, Paris, France; CNRS, UMR 3569, Paris, France; Université Paris Diderot, Sorbonne Paris Cité, Unité de Génétique Moléculaire des Virus à ARN, Paris, France.
authors.forename: Nadia
authors.initials: N
authors.lastname: Naffakh
mesh_terms: D021581:Active Transport, Cell Nucleus; D017398:Alternative Splicing; D002460:Cell Line; D002868:Chromosomal Proteins, Non-Histone / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016207:Cytokines / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D020868:Gene Silencing; D054884:Host-Pathogen Interactions; D006801:Humans; D009980:Influenza A virus / Q000201:enzymology / Q000502:physiology*; D028884:Karyopherins / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D020836:Protein Structure, Quaternary; D012333:RNA, Messenger / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism*; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017381:Spliceosomes / Q000201:enzymology / Q000378:metabolism; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D014779:Virus Replication
chemical_list: D002868:Chromosomal Proteins, Non-Histone; D016207:Cytokines; C090444:IK protein, human; D028884:Karyopherins; C057027:PB2 protein, Influenzavirus A; D012333:RNA, Messenger; D012367:RNA, Viral; D011993:Recombinant Fusion Proteins; C583515:Smu1 protein, human; D014764:Viral Proteins; C077346:influenza virus polymerase basic protein 1; D012324:RNA-Dependent RNA Polymerase