tetano
Editor, Senior Moderator
Virol J. 2018 Mar 27;15(1):55. doi: 10.1186/s12985-018-0960-4.
[h=1]Major contribution of the RNA-binding domain of NS1 in the pathogenicity and replication potential of an avian H7N1 influenza virus in chickens.[/h] Trapp S[SUP]1,[/SUP][SUP]2[/SUP], Soubieux D[SUP]1,[/SUP][SUP]2[/SUP], Lidove A[SUP]1,[/SUP][SUP]2[/SUP], Esnault E[SUP]1,[/SUP][SUP]2[/SUP], Lion A[SUP]1,[/SUP][SUP]2[/SUP], Guillory V[SUP]1,[/SUP][SUP]2[/SUP], Wacquiez A[SUP]1,[/SUP][SUP]2[/SUP], Kut E[SUP]1,[/SUP][SUP]2[/SUP], Qu?r? P[SUP]1,[/SUP][SUP]2[/SUP], Larcher T[SUP]3,[/SUP][SUP]4[/SUP], Ledevin M[SUP]3,[/SUP][SUP]4[/SUP], Nadan V[SUP]5[/SUP], Camus-Bouclainville C[SUP]6[/SUP], Marc D[SUP]7,[/SUP][SUP]8[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]BACKGROUND:[/h] 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.
[h=4]METHODS:[/h] 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.
[h=4]RESULTS:[/h] 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[SUP]- 1[/SUP] 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.
[h=4]CONCLUSIONS:[/h] 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.
[h=4]KEYWORDS:[/h] Chicken; Influenza A; NS1; Viral replication
PMID: 29587792 PMCID: PMC5870492 DOI: 10.1186/s12985-018-0960-4
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[h=1]Major contribution of the RNA-binding domain of NS1 in the pathogenicity and replication potential of an avian H7N1 influenza virus in chickens.[/h] Trapp S[SUP]1,[/SUP][SUP]2[/SUP], Soubieux D[SUP]1,[/SUP][SUP]2[/SUP], Lidove A[SUP]1,[/SUP][SUP]2[/SUP], Esnault E[SUP]1,[/SUP][SUP]2[/SUP], Lion A[SUP]1,[/SUP][SUP]2[/SUP], Guillory V[SUP]1,[/SUP][SUP]2[/SUP], Wacquiez A[SUP]1,[/SUP][SUP]2[/SUP], Kut E[SUP]1,[/SUP][SUP]2[/SUP], Qu?r? P[SUP]1,[/SUP][SUP]2[/SUP], Larcher T[SUP]3,[/SUP][SUP]4[/SUP], Ledevin M[SUP]3,[/SUP][SUP]4[/SUP], Nadan V[SUP]5[/SUP], Camus-Bouclainville C[SUP]6[/SUP], Marc D[SUP]7,[/SUP][SUP]8[/SUP].
[h=3]Author information[/h]
[h=3]Abstract[/h] [h=4]BACKGROUND:[/h] 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.
[h=4]METHODS:[/h] 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.
[h=4]RESULTS:[/h] 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[SUP]- 1[/SUP] 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.
[h=4]CONCLUSIONS:[/h] 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.
[h=4]KEYWORDS:[/h] Chicken; Influenza A; NS1; Viral replication
PMID: 29587792 PMCID: PMC5870492 DOI: 10.1186/s12985-018-0960-4
Free full text