Search

Division of intracellular organelles often correlates with additional membrane wrapping, e.g., by the endoplasmic reticulum or the outer mitochondrial membrane. Such wrapping plays a vital role in proteome and lipidome organization. However, how an extra membrane impacts the mechanics of the division has not been investigated. Here we combine fluorescence and cryo-electron microscopy experiments with self-consistent field theory to explore the stress-induced instabilities imposed by membrane wrapping in a simple double-membrane tubular system. We find that, at physiologically relevant conditions, the outer membrane facilitates an alternative pathway for the inner-tube fission through the formation of a transient contact (hemi-fusion) between both membranes. A detailed molecular theory of the fission pathways in the double membrane system reveals the topological complexity of the process, resulting both in leaky and leakless intermediates, with energies and topologies predicting physiological events.

The mechanochemical GTPase dynamin-related protein 1 (Drp1) catalyzes mitochondrial and peroxisomal fission, but the regulatory mechanisms remain ambiguous. Here we find that a conserved, intrinsically disordered, six-residue Short Linear Motif at the extreme Drp1 C-terminus, named CT-SLiM, constitutes a critical allosteric site that controls Drp1 structure and function in vitro and in vivo. Extension of the CT-SLiM by non-native residues, or its interaction with the protein partner GIPC-1, constrains Drp1 subunit conformational dynamics, alters self-assembly properties, and limits cooperative GTP hydrolysis, surprisingly leading to the fission of model membranes in vitro. In vivo, the involvement of the native CT-SLiM is critical for productive mitochondrial and peroxisomal fission, as both deletion and non-native extension of the CT-SLiM severely impair their progression. Thus, contrary to prevailing models, Drp1-catalyzed membrane fission relies on allosteric communication mediated by the CT-SLiM, deceleration of GTPase activity, and coupled changes in subunit architecture and assembly-disassembly dynamics.

Viruses are very attractive biomaterials owing to their capability as nanocarriers of genetic material. Efforts have been made to functionalize self-assembling viral protein capsids on their exterior or interior to selectively take up different payloads. PRD1 is a double-stranded DNA bacteriophage comprising an icosahedral protein outer capsid and an inner lipidic vesicle. Here, we report the three-dimensional structure of PRD1 in complex with the antipsychotic drug chlorpromazine (CPZ) by cryo-electron microscopy. We show that the jellyrolls of the viral major capsid protein P3, protruding outwards from the capsid shell, serve as scaffolds for loading heterocyclic CPZ molecules. Additional X-ray studies and molecular dynamics simulations show the binding modes and organization of CPZ molecules when complexed with P3 only and onto the virion surface. Collectively, we provide a proof of concept for the possible use of the lattice-like organisation and the quasi-symmetric morphology of virus capsomers for loading heterocyclic drugs with defined properties.

152 p., Viruses are parasitic biological agents that in the virion state contain differentmorphologies and assembly organisations. In some virions, a lipid membrane of cellularorigin is a structural component of the infectious particle. This membrane can belocated as the outermost layer or internally depending on the species. Our work aims atunderstanding the mechanism of assembly and the viral proteins involved in cellularinfection, using for that Cryo-electron microscopy (Cryo-EM) and Cryo-electrontomography (Cryo-ET).In the first case, we present the 3D structures of the archaeal, halophilic, internalmembrane-containing Haloarcula californiae icosahedral virus 1 (HCIV-1) andHaloarcula hispanica icosahedral virus 2 (HHIV-2) at 3.7 and 3.8 Å resolution,respectively, using single-particle cryo-electron microscopy. These structures reveal theviral assembly mechanisms of the capsid shell over the internal membrane and themorphology of the viral receptors.In the second case (ongoing study), using Cryo-ET technique, we have morphologicallycharacterised pleomorphic Bovine Viral Diarrhea Virus 1 (BVDV), as very little isknown about the assembly, virus entry and antibody recognition. Thus, combining cryo-ET with sub-tomogram averaging, we present the preliminary cryo-ET average map ofthe BVDV type 1 bound to mAbs against the glycoproteins Erns.