S T R U C T U R A L B I O L O G Y

S C I E N T I F I C H I G H L I G H T S

4 0 H I G H L I G H T S 2 0 2 1 I

SAM-dependent methyl transferases, enzymes broadly conserved in all kingdoms of life, and that are involved in RNA labelling. The peculiarity of this enzyme is that it is able both to methylate and transfer the CAP structure to the RNA, essential for its stability and recognition by the ribosomes. Below the crown, the complex narrows to the waist, the region defining an internal pore from where the viral RNA could access the interior of viral replication organelles, and from where the newly synthetised viral RNAs would be expelled to the cytoplasm, properly labelled by the capping enzyme. Finally, a twelvefold spiked skirt underneath the waist mediates a very strong interaction with membranes in a monotopic fashion (without traversing the entire lipid bilayer), but reaching a depth of more than 10 Å into the lipid bilayer. The cone shape of the spikes and the surrounding charge distribution are indicative of a very strong potential for membrane deformation. Thus, in its singular architecture, the complex combines the ability to bind and bend membranes, generate a membrane pore and trigger the enzymatic activity, and form a molecular platform for the attachment of other viral factors such as the polymerase [1].

The structure of alphavirus capping rings provides the first high-resolution structural information of the ways RNA viruses replicate and transcribe their genomes in their membranous environment. For example, all alphaviruses (e.g., Semliky forest virus, Venezuelan equine encephalitis

virus), Rubella virus, hepatitis E virus or other related flaviviruses such as Dengue and hepatitis C, or even the coronavirus, carry out replication within membrane organelles. However, the available information about their replication machineries neglects their membrane association, thus current understanding of the way they work limits capacity to, for instance, develop effective antivirals.

This structure makes it possible to better interpret the existing structures of replication complexes at much lower resolution obtained by cryo-electron tomography. This is the case for the structure of the Flock House Virus crown- like structures observed at the necks of spherule-shaped viral organelles [2]. Superposition of this tomographic reconstruction with the nsP1 capping rings illuminates the conservation of these structures between a broad group of positive-stranded RNA viruses (Figure 26). Other tomographic reconstructions of viral factories in coronavirus-infected cells, despite notable differences in the architecture, also show the oligomerisation of replication complexes into macromolecular pores [3]. Thus, the structure of nsP1 capping rings opens a new path for the understanding of viral replication within membrane organelles. This will uncover essential functional aspects of RNA virus biology, possibly leading to the creation of more effective antiviral drugs to contain and fight against future pandemics of emerging infectious diseases by RNA viruses.

Fig. 26: nsP1 complex (coloured by protomer) superposed into the tomography map reconstructed from FHV growing spherules (light grey surface). Left: the complex is shown from the cytosolic view; middle: from the lateral view;

right: from the interior of the viral organelle. The figure was made with ChimeraX.

PRINCIPAL PUBLICATION AND AUTHORS

Capping pores of alphavirus nsP1 gate membranous viral replication factories, R. Jones (a), G. Bragagnolo (a), R. Arranz (b), J. Reguera (a,c), Nature 589, 615-619 (2021); https:/doi.org/10.1038/s41586-020-3036-8 (a) CNRS, AFMB UMR 7257, Aix-Marseille Université, Marseille (France) (b) National Center of Biotechnology, CSIC, Madrid (Spain) (c) INSERM, AFMB UMR7257, Marseille (France)

REFERENCES

[1] M.K. Pietila, K. Hellstrom, T. Ahola, Virus Research 234, 44-57 (2017). [2] N. Unchwaniwala et al., PNAS 117, 18680-18691 (2020). [3] G. Wolff et al., Science 369, 1395-1398 (2020).