Projets de recherche
Interactions between avian and influenza virus proteins
Project funded by a Marie Curie Career Integration Grant (2013-2017) to Mariette Ducatez
Influenza virus is a segmented single stranded negative sense RNA virus of the Orthomyxoviridae family, which includes three Genera: influenza A, B, and C . We will focus on influenza A virus and study host/pathogen interactions. Influenza A strains are classified into subgroups (or serotypes) based on their surface glycoproteins: hemagglutinin (HA) and neuraminidase (NA). Today we know 16 HA and 9 NA types. Aquatic birds are the natural reservoir of influenza A viruses and many mammals can also be infected by the virus either sporadically or with well adapted lineages. Human, swine and horse are for instance natural hosts of H1N1 and H3N2; H1N1, H1N2, and H3N2; H3N8 and H7N7 strains, respectively .
On rare occasions, swine influenza viruses can infect humans (and human viruses infect swine), generally resulting in a mild disease [2, 3]. Avian influenza viruses (AIVs) can also sporadically infect humans. It occurred in 1999 and 2003 in Hong Kong following H9N2 virus cases in poultry [4-6], in 2003 in the Netherlands after a large outbreak of highly pathogenic avian influenza (HPAI) H7N7 virus in poultry , or since 2003 in Asia and Africa with HPAI H5N1 virus.
Avian influenza interspecies transmissions are rare but remain a major concern for public health.
Avian influenza was indeed involved in all influenza pandemics identified so far. The 1918 H1N1 “Spanish flu” pandemic origin is still debated and was caused either by a direct zoonotic transmission from avian to human or by a re-assortment between avian, swine and human gene segments in human. Similarly Asian H2N2 (1957-58) and Hong Kong H3N2 (1968-69) pandemics also emerged after the introduction of avian influenza gene segments .
State of the art
The systematic study of interactions between the viral protein and host cells is a powerful approach for characterizing the determinants of viral cellular infection.
In influenza both the non structural protein 1 (NS1) and the ribonucleoproteic complex proteins have been shown to widely interact with host cellular proteins on a mammal model.
Interactions between NS1 and cellular proteins
NS1 has been identified as an important virulence factor of the influenza virus. Multiple functions have been attributed to NS1 as reviewed by Hale et al. in 2008 . NS1 mainly impacts the host immune response: it reduces the induction of IFN-β , inhibits the function of intracytoplasmic pathogen sensor human retinoic acid-inducible gene product I (RIG-I) , and acts on the host protein synthesis by suppressing the protein kinase (PKR) mediated inhibition of replication  and limiting the activation of the 2′-5′-oligo (A) synthetase/RNase L pathway . NS1 together with phosphatidylinositol-3-kinase (PI3K) can prevent apoptosis during infection .
In the last couple of years, several research groups have worked on the interactions between NS1 and cellular proteins. In mammal cells (293T, A549, and/or HeLa cells), they have shown that NS1 interacts with Scribble, MAGI-1,2,3, Dgl1 [15, 16], Akt(-pH) , Pdlim2 , Hsp90 , hnRNP-F , hStau-1 , RIL, regulating cellular tyrosine kinase Src (Bavagnoli et al., 2011), and PSD-95 (Zhang et al., 2011) as shown by pull-down, two-hybrid and immunofluorescence assays and sometimes confirmed by interference RNA experiments. Thanks to RNAseq and microarray technologies, NS1 has also been shown to bind to hPAF1C (provided NS1 possessed a histone-like sequence), resulting in the suppression of hPAF1C-mediated transcriptional elongation and with consequences on the antiviral response (Marazzi et al., 2012). The role of the discovered interactions is not always well understood yet. MAGI proteins play a role in the membrane permeability , and Dgl1 and Scribble associated with NS1 disrupt tight junctions (Gobiewski et al., 2011). HnRNP-F and hStau-1 with NS1 seem to increase viral replication [20, 21]. Several interactions moreover suggest a regulation (or even often an inhibition) of virus-induced apoptosis such as Scribble, Akt, Hsp90 [15, 17, 19].
Avian and mammal NS1 C-terminal motifs differ with ESEV and RSKV on avian and mammal NS1, respectively. These C-terminal motifs have also been associated with interspecies transmission as well as with species-specific virulence [22, 23]. The anti-apoptotic effect of NS1-Scribble interaction was shown with the avian NS1 ESEV motif . The nuclear localization of NS1 in mammal cells (but not avian cells) seem to also be dependent on the amino acid (aa) sequence of the C-terminal part of NS1 .
Objectives of the proposal and expected results
To date most of the knowledge on influenza virus and host cell interactions was gained using mammal models. Very little is known on avian cell/influenza virus interactions. Both the influenza non structural protein 1 (NS1) and the ribonucleoproteic complex proteins have been shown to play an important role during the host cell infection process. However, there are well established differences between avian and mammal NS1 and avian and mammal hosts (cells) also display obvious specificities.
We therefore propose to study the interactions between influenza virus and host proteins in an avian model and to compare them with what is known on the mammalian model.
Our project has three operational objectives:
- Objective 1: identify the avian cellular proteins that interact with influenza virus proteins
- Objective 2: narrow the identified interaction(s) to the protein domain level and identify molecular determinants of host specificity
- Objective 3 (longer term): study the mechanism of interaction
We are expecting to observe both similarities and differences in the interactions between influenza proteins and avian cells and those reported in mammal cells. This comparison could allow us to identify proteins or protein domains, which play an important role in interspecies transmission.
Our final goal is to highlight parts of the virus genome to closely survey for pandemic preparedness.
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21. de Lucas, S., et al., Human Staufen1 protein interacts with influenza virus ribonucleoproteins and is required for efficient virus multiplication. J Virol, 2010. 84(15): p. 7603-12.
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