A first article published in 2023 proposed an initial model for the encapsidation of Influenza virus genome. In a new publication published in the Nucleic Acids Research journal, the same scientists have now obtained a high-resolution structure of the Influenza nucleoprotein in an antiparallel helix that now details precisely all the protein-protein and protein-RNA interactions within the nucleocapsid.
Every year, Influenza virus causes major epidemics affecting between 2 to 6 million people in France. The strains responsible for these epidemics are very similar to viruses that infect other animal species, giving rise to constant fears of new emerging pandemic strains. In this context, the actual circulation of the avian Influenza virus in cattle farms in the US is being closely monitored.
The genome of Influenza A virus is made up of eight single-stranded RNA molecules of negative polarity. Each RNA segment is encapsidated by multiple copies of the viral nucleoprotein (NPs) with their 3‘ and 5’ ends interacting with an RNA polymerase to form the ribonucleoprotein complex (RNP), a functional entity in viral proliferation. Extracted directly from the virus and observed by electron microscopy (A), RNPs appear to be intricate antiparallel double-stranded helices that are extremely flexible and highly dynamic. Their structural studies by cryo-electron microscopy (cryo-EM) had merely suggested an imprecise positioning of the NP molecules, without providing any details of the precise interaction with the viral RNA. In 2023, scientists from the Institut de Biologie Structurale (IBS) in Grenoble published an article in Science Advances to present a method to assemble nucleocapsid-like particles in vitro from recombinant NP and short RNAs. They had obtained a first parallel double-stranded helical model. This model suffered from several limitations, in particular regarding the direction of the two helical strands and the precision of the protein-RNA interactions to stabilize the integrity of the whole structure.
By slightly optimising the protein sequence of NP, the scientists succeeded in strengthening the integrity of the nucleocapsid-like particles, and the Glacios microscope of the ISBG electron microscopy platform (UMS3518) enabled them to obtain a high-resolution 3D reconstruction of the parallel double-stranded helix. This strategy also opened the way to other improvements, in particular in terms of the length of the RNA probes used. By increasing the length of the RNAs, they obtained several 3D reconstructions at the atomic-scale, including one with the helices arranged in anti-parallel conformation (B). The scientists now have a whole series of high-resolution 3D reconstructions to understand how NP can arrange into a helix. In particular, the antiparallel reconstruction shows how the two helices interact within this flexible architecture. They also know for the first time how the RNA roles over the entire surface of NP (C). The models show that while RNA is involved in structuring the helix, it can adopt different conformations. These data also pave the way for the design of molecules that could specifically target the protein-RNA interaction of NP, to prevent its interaction with the viral RNA, thereby limiting the formation of RNPs and preventing the proliferation of Influenza virus.
Influenza A virus antiparallel helical nucleocapsid-like pseudo-atomic structure. Florian Chenavier, Eleftherios Zarkadas, Lily-Lorette Freslon, Alice J. Stelfox, Guy Schoehn, Rob W.H. Ruigrok, Allison Ballandras-Colas, Thibaut Crépin. Nucleic Acids Research 2024 Dec 14:gkae1211. doi : 10.1093/nar/gkae1211.
Contact : Thibaut Crepin (IBS/Viral Replication Machines Group)