Major contribution of the RNA-binding domain of NS1 in the pathogenicity and replication potential of an avian H7N1 influenza virus in chickens

作者：Trapp Sascha; Soubieux Denis; Lidove Alexandra; Esnault Evelyne; Lion Adrien; Guillory Vanaique; Wacquiez Alan; Kut Emmanuel; Quere Pascale; Larcher Thibaut; Ledevin Mireille; Nadan Virginie; Camus Bouclainville Christelle; Marc Daniel^*

来源：Virology Journal, 2018, 15(1): 55.

DOI：10.1186/s12985-018-0960-4

• Influenza A
• NS1
• Viral replication
• Chicken

Background: Non-structural protein NS1 of influenza A viruses harbours several determinants of pathogenicity and host-range. However it is still unclear to what extent each of its two structured domains (i.e. RNA-binding domain, RBD, and effector domain, ED) contribute to its various activities.
Methods: To evaluate the respective contributions of the two domains, we genetically engineered two variants of an H7N1 low pathogenicity avian influenza virus harbouring amino-acid substitutions that impair the functionality of either domain. The RBD- and ED-mutant viruses were compared to their wt-counterpart in vivo and in vitro, notably in chicken infection and avian cell culture models.
Results: The double substitution R38A-K41A in the RBD dramatically reduced the pathogenicity and replication potential of the virus, whereas the substitution A149V that was considered to abrogate the IFN-antagonistic activity of the effector domain entailed much less effects. While all three viruses initiated the viral life cycle in avian cells, replication of the R38A-K41A virus was severely impaired. This defect was associated with a delayed synthesis of nucleoprotein NP and a reduced accumulation of NS1, which was found to reach a concentration of about 30 micromol. L-1 in wt-infected cells at 8 h post-infection. When overexpressed in avian lung epithelial cells, both the wt-NS1 and 3841AA-NS1, but not the A149V-NS1, reduced the poly(I:C)-induced activation of the IFN-sensitive chicken Mx promoter. Unexpectedly, the R38A-K41A substitution in the recombinant RBD did not alter its in vitro affinity for a model dsRNA. When overexpressed in avian cells, both the wt- and A149V-NS1s, as well as the individually expressed wt-RBD to a lesser extent, enhanced the activity of the reconstituted viral RNA-polymerase in a minireplicon assay.
Conclusions: Collectively, our data emphasized the critical importance and essential role of the RNA-binding domain in essential steps of the virus replication cycle, notably expression and translation of viral mRNAs.

• 出版日期2018-3-27