Abstract
The IFN-induced human myxovirus resistance protein A (MxA) exhibits a broad antiviral activity against many viruses, including influenza A virus (IAV). MxA belongs to the family of dynamin-like GTPases and assembles in vitro into dimers, tetramers, and oligomeric ring-like structures. The molecular mechanism of action remains to be elucidated. Furthermore, it is not clear whether MxA exerts its antiviral activity in a monomeric and/or multimeric form. Using a set of MxA mutants that form complexes with defined stoichiometry, we observed that, in the presence of guanosine 5'-O-(thiotriphosphate), purified MxA disassembled into tetramers and dimers. Dimeric forms did not further disassemble into monomers. Infection experiments revealed that besides wild-type MxA, dimeric and monomeric variants of MxA also efficiently restricted IAV at a replication step after primary transcription. Moreover, only dimeric MxA was able to form stable complexes with the nucleoprotein (NP) of IAV. MxA interacted with NP independently of other viral components. Interestingly, the dimeric form of MxA was able to efficiently bind to NP from several MxA-sensitive strains but interacted much more weakly with NP from the MxA-resistant PR8 strain derived from the H1N1 1918 lineage. Taken together, these data suggest that, during infection, a fraction of MxA disassembles into dimers that bind to NP synthesized following primary transcription in the cytoplasm, thereby preventing viral replication.
Highlights
Proteins are members of the dynamin superfamily of large GTPases
In the case of influenza A virus (IAV), several previous studies have demonstrated that ectopic expression of the nucleocapsid protein (NP) or the polymerase subunit 2 (PB2) of the virus partially abrogated the antiviral activity of mouse Mx1 [15,16,17]
Oligomeric myxovirus resistance protein A (MxA) Disassembles upon GTP Binding into Tetramers and Dimers—To investigate whether MxA has to first adopt a defined oligomeric or monomeric form to exert antiviral activity against IAV, we made use of several mutants of MxA that form complexes with a defined stoichiometry (Fig. 1A) [5, 12]
Summary
Proteins are members of the dynamin superfamily of large GTPases. They contain an amino-terminal globular GTPase (G) domain and a carboxy-terminal stalk domain [5, 6]. We transfected HEK293T cells with plasmids encoding wild-type MxA, the interface variants MxA(L617D) (dimer) and MxA(M527D) (monomer), and the hinge 1 variant MxA(R640A) (dimer/tetramer) as well as the inactive MxA(T103A) and infected them with the MxA-sensitive IAV strain pH1N1-NP (H5N1) at various concentrations [18].
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