More Like this

1. From Disorder to Icosahedral Symmetry: How Conformation-Switching Subunits Enable RNA Virus Assembly

   Li, S; Tresset, G; Zandi, R (August 2025, Science advances)

   Icosahedral capsids are ubiquitous among spherical viruses, yet their assem- bly pathways and governing interactions remain elusive. We present a molecular dynamics model that incorporates essential physical and biological interactions, including protein diffusion, genome flexibility, and a conformational switch that mimics allostery and activates the elastic properties of proteins upon binding. This switch makes the simulations computationally feasible and enables the assembly of icosahedral capsids around a flexible genome—overcoming long-standing lim- itations in previous models. Using this framework, we successfully reproduce the self-assembly of subunits around a flexible genome into icosahedral shells with numbers greater than one – most notably 3, the most common structure in na- ture – a feat that rigid-body models have so far failed to achieve. We systematically explore the range of morphologies formed with different genome architectures, in line with in vitro experiments using cowpea chlorotic mottle virus capsid proteins: viral RNAs with more complex structure form more complete and stable capsids than linear ones. These results provide a predictive framework for genome-guided assembly and capsid design. 

   more » « less
2. Phosphatidylinositol-3-phosphate mediates Arc capsid secretion through the multivesicular body pathway

   https://doi.org/10.1073/pnas.2322422121  

   Mehta, Kritika; Yentsch, Henry; Lee, Jungbin; Yook, Yeeun; Lee, Kwan Young; Gao, Tianyu Terry; Tsai, Nien-Pei; Zhang, Kai (August 2024, Proceedings of the National Academy of Sciences)

   Activity-regulated cytoskeleton-associated protein (Arc/Arg3.1) is an immediate early gene that plays a vital role in learning and memory. Arc protein has structural and functional properties similar to viral Group-specific antigen (Gag) protein and mediates the intercellular RNA transfer through virus-like capsids. However, the regulators and secretion pathway through which Arc capsids maneuver cargos are unclear. Here, we identified that phosphatidylinositol-3-phosphate (PI3P) mediates Arc capsid assembly and secretion through the endosomal–multivesicular body (MVB) pathway. Indeed, reconstituted Arc protein preferably binds to PI3P. In HEK293T cells, Arc forms puncta that colocalize with FYVE, an endosomal PI3P marker, as well as Rab5 and CD63, early endosomal and MVB markers, respectively. Superresolution imaging resolves Arc accumulates within the intraluminal vesicles of MVB. CRISPR double knockout of RalA and RalB, crucial GTPases for MVB biogenesis and exocytosis, severely reduces the Arc-mediated RNA transfer efficiency. RalA/B double knockdown in cultured rat cortical neurons increases the percentage of mature dendritic spines. Intake of extracellular vesicles purified from Arc-expressing wild-type, but not RalA/B double knockdown, cells in mouse cortical neurons reduces their surface GlutA1 levels. These results suggest that unlike the HIV Gag, whose membrane targeting requires interaction with plasma-membrane-specific phosphatidyl inositol (4,5) bisphosphate (PI(4,5)P2), the assembly of Arc capsids is mediated by PI3P at endocytic membranes. Understanding Arc’s secretion pathway helps gain insights into its role in intercellular cargo transfer and highlights the commonality and distinction of trafficking mechanisms between structurally resembled capsid proteins. 

   more » « less
3. The In Vitro Packaging of “Overlong” RNA by Spherical Virus-Like Particles

   Duran-Meza, A L; Chapman, A G; Tanimoto, C R; Knobler, C M; Gelbart, W M (November 2023, Physical VIrology Springer Series in Biophysics)

   Of the myriad viruses, very few have been shown to be capable of self- assembly in vitro from purified components into infectious virus particles. One of these is Cowpea Chlorotic Mottle Virus (CCMV), an unenveloped spherical plant virus whose capsid self-assembles around its RNA genome without a packaging signal. While heterologous RNA, not just cognate viral RNA, can be packaged into individual CCMV virus-like particles (VLPs), the RNA needs to fall within a certain range of lengths. If it is too short, it is packaged into particles smaller than wild type, or with two or more RNAs per capsid. If the RNA is too long, multiple capsids assemble around one RNA, and the RNA associated with these multiplet structures is not as RNase resistant. Further, as shown in the present work, 4200 nt appears to be the limiting length of RNA that can be packaged into single RNase-resistant CCMV VLPs. We explore the extent to which “overlong” RNA can be packaged more efficiently upon the addition of spermine, a polyvalent cation whose increasing concentration has been shown to compactify RNA. Finally, we show that the capsid protein of Brome Mosaic Virus (BMV), a bromovirus closely related to CCMV, also gives rise to multiplets when it is self-assembled with the same “overlong” RNA constructs, but with different distributions of multiplets. 

   more » « less
4. Genome organization and interaction with capsid protein in a multipartite RNA virus

   https://doi.org/10.1073/pnas.1915078117  

   Beren, Christian; Cui, Yanxiang; Chakravarty, Antara; Yang, Xue; Rao, A. L.; Knobler, Charles M.; Zhou, Z. Hong; Gelbart, William M. (May 2020, Proceedings of the National Academy of Sciences)

   null (Ed.)

   We report the asymmetric reconstruction of the single-stranded RNA (ssRNA) content in one of the three otherwise identical virions of a multipartite RNA virus, brome mosaic virus (BMV). We exploit a sample consisting exclusively of particles with the same RNA content—specifically, RNAs 3 and 4—assembled in planta by agrobacterium-mediated transient expression. We find that the interior of the particle is nearly empty, with most of the RNA genome situated at the capsid shell. However, this density is disordered in the sense that the RNA is not associated with any particular structure but rather, with an ensemble of secondary/tertiary structures that interact with the capsid protein. Our results illustrate a fundamental difference between the ssRNA organization in the multipartite BMV viral capsid and the monopartite bacteriophages MS2 and Qβ for which a dominant RNA conformation is found inside the assembled viral capsids, with RNA density conserved even at the center of the particle. This can be understood in the context of the differing demands on their respective lifecycles: BMV must package separately each of several different RNA molecules and has been shown to replicate and package them in isolated, membrane-bound, cytoplasmic complexes, whereas the bacteriophages exploit sequence-specific “packaging signals” throughout the viral RNA to package their monopartite genomes. 

   more » « less
5. A computational spatial whole-Cell model for hepatitis B viral infection and drug interactions

   https://doi.org/10.1038/s41598-023-45998-0  

   Ghaemi, Zhaleh; Nafiu, Oluwadara; Tajkhorshid, Emad; Gruebele, Martin; Hu, Jianming (December 2023, Scientific Reports)

   Abstract Despite a vaccine, hepatitis B virus (HBV) remains a world-wide source of infections and deaths. We develop a whole-cell computational platform combining spatial and kinetic models describing the infection cycle of HBV in a hepatocyte host. We simulate key parts of the infection cycle with this whole-cell platform for 10 min of biological time, to predict infection progression, map out virus-host and virus-drug interactions. We find that starting from an established infection, decreasing the copy number of the viral envelope proteins shifts the dominant infection pathway from capsid secretion to re-importing the capsids into the nucleus, resulting in more nuclear-localized viral covalently closed circular DNA (cccDNA) and boosting transcription. This scenario can mimic the consequence of drugs designed to manipulate viral gene expression. Mutating capsid proteins facilitates capsid destabilization and disassembly at nuclear pore complexes, resulting in an increase in cccDNA copy number. However, excessive destabilization leads to premature cytoplasmic disassembly and does not increase the cccDNA counts. Finally, our simulations can predict the best drug dosage and its administration timing to reduce the cccDNA counts. Our adaptable computational platform can be parameterized to study other viruses and identify the most central viral pathways that can be targeted by drugs. 

   more » « less