title: Identification of putative interactions between swine and human influenza A virus nucleoprotein and human host proteins.
abstract: BACKGROUND
  Influenza A viruses (IAVs) are important pathogens that affect the health of humans and many additional animal species. IAVs are enveloped, negative single-stranded RNA viruses whose genome encodes at least ten proteins. The IAV nucleoprotein (NP) is a structural protein that associates with the viral RNA and is essential for virus replication. Understanding how IAVs interact with host proteins is essential for elucidating all of the required processes for viral replication, restrictions in species host range, and potential targets for antiviral therapies.
  
  
  METHODS
  In this study, the NP from a swine IAV was cloned into a yeast two-hybrid "bait" vector for expression of a yeast Gal4 binding domain (BD)-NP fusion protein. This "bait" was used to screen a Y2H human HeLa cell "prey" library which consisted of human proteins fused to the Gal4 protein's activation domain (AD). The interaction of "bait" and "prey" proteins resulted in activation of reporter genes.
  
  
  RESULTS
  Seventeen positive bait-prey interactions were isolated in yeast. All of the "prey" isolated also interact in yeast with a NP "bait" cloned from a human IAV strain. Isolation and sequence analysis of the cDNAs encoding the human prey proteins revealed ten different human proteins. These host proteins are involved in various host cell processes and structures, including purine biosynthesis (PAICS), metabolism (ACOT13), proteasome (PA28B), DNA-binding (MSANTD3), cytoskeleton (CKAP5), potassium channel formation (KCTD9), zinc transporter function (SLC30A9), Na+/K+ ATPase function (ATP1B1), and RNA splicing (TRA2B).
  
  
  CONCLUSIONS
  Ten human proteins were identified as interacting with IAV NP in a Y2H screen. Some of these human proteins were reported in previous screens aimed at elucidating host proteins relevant to specific viral life cycle processes such as replication. This study extends previous findings by suggesting a mechanism by which these host proteins associate with the IAV, i.e., physical interaction with NP. Furthermore, this study revealed novel host protein-NP interactions in yeast.
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. generous.alex@mayo.edu.
authors.forename: Alex
authors.initials: A
authors.lastname: Generous
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. mttho@umich.edu.
authors.forename: Molly
authors.initials: M
authors.lastname: Thorson
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. barcu010@umn.edu.
authors.forename: Jeff
authors.initials: J
authors.lastname: Barcus
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. joejacher@gmail.com.
authors.forename: Joseph
authors.initials: J
authors.lastname: Jacher
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. marc.busch@drake.edu.
authors.forename: Marc
authors.initials: M
authors.lastname: Busch
authors.affiliation: Biology Department, Drake University, 1344 27th St., Des Moines, IA, 50311, USA. heidi.sleister@drake.edu.
authors.forename: Heidi
authors.initials: H
authors.lastname: Sleister
mesh_terms: D000818:Animals; D017930:Genes, Reporter; D006367:HeLa Cells; D054884:Host-Pathogen Interactions; D006801:Humans; D029521:Alphainfluenzavirus / Q000302:isolation & purification / Q000502:physiology*; D019590:Nucleocapsid Proteins; D025941:Protein Interaction Mapping; D016601:RNA-Binding Proteins / Q000378:metabolism*; D013552:Swine; D020798:Two-Hybrid System Techniques; D014758:Viral Core Proteins / Q000378:metabolism*
chemical_list: C048842:NP protein, Influenza A virus; D019590:Nucleocapsid Proteins; D016601:RNA-Binding Proteins; D014758:Viral Core Proteins