Nanoscale organization of rotavirus replication machineries

Abstract

Rotavirus genome replication and assembly take place in cytoplasmic electron dense inclusions termed viroplasms (VPs). Previous conventional optical microscopy studies observing the intracellular distribution of rotavirus proteins and their organization in VPs have lacked molecular-scale spatial resolution, due to inherent spatial resolution constraints. In this work we employed super-resolution microscopy to reveal the nanometric-scale organization of VPs formed during rotavirus infection, and quantitatively describe the structural organization of seven viral proteins within and around the VPs. The observed viral components are spatially organized as five concentric layers, in which NSP5 localizes at the center of the VPs, surrounded by a layer of NSP2 and NSP4 proteins, followed by an intermediate zone comprised of the VP1, VP2, VP6. In the outermost zone, we observed a ring of VP4 and finally a layer of VP7. These findings show that rotavirus VPs are highly organized organelles.

Coding-Gene Coevolution Analysis of Rotavirus Proteins: A Bioinformatics and Statistical Approach

“…Although coevolving residues between VP1 and VP3/VP2 are expected, the finding that VP1 residues coevolve also with NSP4 was unexpected, since it is known that NSP4 functions as an intracellular receptor in the endoplasmic reticulum membrane (ER) [85] that may play an important role in virion assembly at the late stages of viral replication according to different hypotheses [86][87][88]. However, it was reported recently that NSP4 can accumulate in proximity of viral replication components, mainly NSP2, NSP5, VP1, VP2, and VP6 [89], suggesting that it also plays an important role as a regulator of viral particle assembly. In addition, intra-co-evolving residues within VP1 were located to the N-and C-terminal of the protein, as supported by a recent report showing that N-and C-terminal domains of VP1 regulate its function [90].…”

Towards a Quantitative Single Particle Characterization by Super Resolution Microscopy: From Virus Structures to Antivirals Design

“…(C) Rotavirus SRM 3B images of different proteins of the viroplasm. Right: model of distribution of viral components ( Garcés Suárez et al, 2019 ).…”

“…The structure of the Rotavirus, a dsRNA virus that mainly produces gastroenteritis disease, was also characterized using a SRM technique called “Bayesian Blinking and Bleaching” (3B). Garcés Suárez et al (2019) analyzed the relative distribution of seven different proteins in the viroplasm of the rotavirus, and developed a model with the spatial distribution of all the proteins in the virosomes. Using the model, they observed the arrangement of the seven proteins in five different concentric layers with highly organized organelles ( Figure 2C ) ( Garcés Suárez et al, 2019 ).…”

Advances in the Development of Antiviral Compounds for Rotavirus Infections

“…The formation of complexes between VPs and LDs is crucial for RV replication, since compounds disrupting LDs significantly decrease the numbers and sizes of VPs and the RV yield (78,102). In a recent study, Suárez et al proposed a model of the nanometric-scale organization of VPs (103). The authors showed that the viral components in the VPs form porous circumferences that are arranged as quite discrete concentric layers.…”