The CPSF30 binding site on the NS1A protein of influenza A virus is a potential antiviral target.
The emergence of influenza A viruses resistant to the two existing classes of antiviral drugs highlights the need for additional antiviral drugs, particularly considering the potential threat of a pandemic of H5N1 influenza A viruses. Here, we determine whether influenza A virus replication can be selectively inhibited by blocking the ability of its NS1A protein to inhibit the 3'-end processing of cellular pre-mRNAs, including beta interferon (IFN-beta) pre-mRNA. Pre-mRNA processing is inhibited via the binding of the NS1A protein to the cellular CPSF30 protein, and mutational inactivation of this NS1A binding site causes severe attenuation of the virus. We demonstrate that binding of CPSF30 is mediated by two of its zinc fingers, F2F3, and that the CPSF30/F2F3 binding site on the NS1A protein extends from amino acid 144 to amino acid 186. We generated MDCK cells that constitutively express epitope-tagged F2F3 in the nucleus, although at only approximately one-eighth the level of the NS1A protein produced during virus infection. Influenza A virus replication was inhibited in this cell line, whereas no inhibition was observed with influenza B virus, whose NS1B protein lacks a binding site for CPSF30. Influenza A virus, but not influenza B virus, induced increased production of IFN-beta mRNA in the F2F3-expressing cells. These results, which indicate that F2F3 inhibits influenza A virus replication by blocking the binding of endogenous CPSF30 to the NS1A protein, point to this NS1A binding site as a potential target for the development of antivirals directed against influenza A virus.
Institute for Cellular and Molecular Biology, Section of Molecular Genetics and Microbiology, University of Texas at Austin, 2500 Speedway, Austin, Texas 78712, USA.
Karen Y
KY
Twu
Diana L
DL
Noah
Ping
P
Rao
Rei-Lin
RL
Kuo
Robert M
RM
Krug
D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D001665:Binding Sites; D002460:Cell Line; D039223:Cleavage And Polyadenylation Specificity Factor / Q000378:metabolism*; D004285:Dogs; D006801:Humans; D009980:Influenza A virus / Q000187:drug effects / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology*; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000378:metabolism*; D014779:Virus Replication / Q000187:drug effects*; D016335:Zinc Fingers
D000998:Antiviral Agents; C000594014:CPSF4 protein, human; D039223:Cleavage And Polyadenylation Specificity Factor; C085820:INS1 protein, influenza virus; D010446:Peptide Fragments; D017361:Viral Nonstructural Proteins