Replication and transcription of the bunyavirus genome

The viral order Bunyavirales contains 450 viruses grouped into 12 families. Among them are several pathogens such as the Hantaan virus, which causes haemorrhagic fevers in humans, or the La Crosse virus, which causes encephalitis in newborns and young children. Our aim is to observe and understand how the components of these viruses work at the molecular level.

This is interesting in more than one way: we can visualise with cryo-electron microscopy machineries present in these viruses at a quasi-atomic resolution and thus understand with very fine details their mode of action. This may also be important for the future development of anti-virals that block certain essential steps in the viral cycle.

We are focusing on two key steps in the viral cycle: replication of the RNA genome of these viruses, which copies the genetic material, and transcription, which produces messenger RNAs that are translated into viral proteins. The neo-synthesised proteins and RNAs will be essential for the formation of new virions.

The viral polymerase is the main enzyme in charge of replication and transcription. A true multifunctional machine, this large protein (250kDa) modifies its organization to successfully perform all its functions. We express it, purify it, characterise its activity and seek to capture its structure in different conformations in order to reconstruct a molecular film of its activity.

The substrate of the polymerase is RNA which is coated with nucleoproteins. The assembly thus formed, called nucleocapsid, is also our subject of study. We seek to understand how nucleoproteins protect the RNA from the environment while allowing the polymerase access for genome replication and transcription.

In the future we will seek to understand at the molecular level how the three players, polymerase, RNA and nucleoproteins interact with each other to form the ribonucleoprotein complex (RNP) and thus perform replication and transcription in a coordinated manner. We will also analyse the interactions of these partners with proteins in the infected cell.
In terms of methods, we use biochemistry to purify and measure the activity of our proteins and cryo-electron microscopy of isolated particles to determine their structures. In the future we will couple this with cryo-electron tomography. For this we use the cryo-microscopes of the platform maintained by the group as well as the Titan Krios at the ESRF.

Main publications :
1. Arragain B, Durieux Trouilleton Q Baudin F, Provaznik J, Azevedo N, Cusack S*, Schoehn G, Malet H* Nature Communications, 2022 Feb 16;13(1):902. Structural snapshots of La Crosse virus polymerase reveal the mechanisms underlying Peribunyaviridae replication and transcription.
2. Arragain B, Effantin G, Gerlach P, Reguera J, Schoehn G, Cusack S*, Malet H*. Nature Communications. 2020 Jul 17;11(1):3590. Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes.
3. Arragain B, Reguera J, Desfosses A, Gutsche I, Schoehn G*, Malet H*. eLife 2019;8:e43075 High resolution cryo-EM structure of the helical RNA-bound Hantaan virus nucleocapsid reveals its assembly mechanisms.