CASC3 Biomolecular Condensates Restrict Turnip Crinkle Virus by Limiting Host Factor Availability

Rademacher, Dana J; Bello, Abudu I.; May, Jared

doi:10.1016/j.jmb.2023.167956

Cited by 6 publications

(3 citation statements)

References 79 publications

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“…Sequestration of cytosolic enzymes by vir-condensates likely occurs in direct competition with cellular condensates that sequester these same enzymes (e.g., GAPDH). A recent study found that the antiviral CASC3 condensates efficiently sequester tombusvirus host factors like GAPDH, Vps29, and HSP70 [67].…”

Section: Discussionmentioning

confidence: 99%

Co-opted cytosolic host proteins form unique condensate substructures within the membranous tombusviral replication organelles

Lin¹,

Nagy²

2023

Preprint

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Positive-strand RNA viruses co-opt intracellular organellar membranes for the biogenesis of viral replication organelles (VROs). The membranous VROs are the sites of viral replication. Tombusviruses co-opt numerous host proteins, many of them cytosolic, not membrane-bound, such as the glycolytic and fermentation enzymes. It is currently not known what type of molecular organization keeps these enzymes enriched and sequestered within the membranous VROs. Here, we show evidence that the tomato bushy stunt virus (TBSV) p33 and the closely-related carnation Italian ringspot virus (CIRV) p36 replication proteins sequester several co-opted cytosolic proteins in unique condensate substructures associated with the membranous VROs. We find that TBSV p33 and CIRV p36 replication proteins form dropletsin vitroorganized by their intrinsically disordered region (IDR). The replication proteins also organize the partitioning of several co-opted host proteins to p33/p36 droplets. We also show that the VRO-associated condensates containing co-opted glycolytic and fermentation enzymes are critical for local ATP production within the VRO to support the energy need of virus replication. A short segment with charged amino acids in p33/p36 IDR plays a critical role in droplet formationin vitro, in organizing the VRO-associated condensates and affects both ATP production in VROs and viral replication. We find that the co-opted ER membranes and actin filaments form meshworks within and around the condensate, likely contributing to the unique composition and structure of VROs. We propose that the p33/p36 replication proteins organize liquid-liquid phase separation of co-opted highly concentrated host proteins in condensate substructures within VROs to allow for the dynamic regulation of viral replication. Overall, we demonstrate that distinct substructures co-exist in tombusvirus VROs. These are the subverted membranes and condensate substructures, both of which are critical for VRO functions. The p33/p36 replication proteins induce and connect the two substructures within the VROs. Interfering with condensate formation in VROs might open up new antiviral approaches.

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Section: Discussionmentioning

confidence: 99%

Co-opted cytosolic host proteins form unique condensate substructures within the membranous tombusviral replication organelles

Lin¹,

Nagy²

2023

Preprint

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“…GAPDH). A recent study found that the antiviral CASC3 condensates efficiently to sequester tombusvirus host factors such as GAPDH, Vps29, and HSP70 (Rademacher et al ., 2023).…”

Section: Discussionmentioning

confidence: 99%

Co‐opted cytosolic proteins form condensate substructures within membranous replication organelles of a positive‐strand RNA virus

Lin,

Nagy

2024

New Phytologist

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Summary Positive‐strand RNA viruses co‐opt organellar membranes for biogenesis of viral replication organelles (VROs). Tombusviruses also co‐opt pro‐viral cytosolic proteins to VROs. It is currently not known what type of molecular organization keeps co‐opted proteins sequestered within membranous VROs. In this study, we employed tomato bushy stunt virus (TBSV) and carnation Italian ringspot virus (CIRV) – Nicotiana benthamiana pathosystems to identify biomolecular condensate formation in VROs. We show that TBSV p33 and the CIRV p36 replication proteins sequester glycolytic and fermentation enzymes in unique condensate substructures associated with membranous VROs. We find that p33 and p36 form droplets in vitro driven by intrinsically disordered region. The replication protein organizes partitioning of co‐opted host proteins into droplets. VRO‐associated condensates are critical for local adenosine triphosphate production to support energy for virus replication. We find that co‐opted endoplasmic reticulum membranes and actin filaments form meshworks within and around VRO condensates, contributing to unique composition and structure. We propose that p33/p36 organize liquid–liquid phase separation of co‐opted concentrated host proteins in condensate substructures within membranous VROs. Overall, we demonstrate that subverted membranes and condensate substructures co‐exist and are critical for VRO functions. The replication proteins induce and connect the two substructures within VROs.

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“…As described above, sequestering pro‐viral factors in viral condensates can enhance virus fitness, but it is also conceivable that host condensates could sequester these same factors and limit their availability to viruses. In support of this, sequestration of pro‐viral host factors like heat shock protein 70 (HSP70) and GAPDH into cellular CASC3 condensates, severely restricts Turnip crinkle virus (TCV, Tombusviridae family) accumulation in Nicotiana benthamiana (Rademacher et al ., 2023). Thus, further elucidating the ‘interactomes’ of both viral and host condensates will likely reveal significant competition for specific biomolecules.…”

Section: Figmentioning

confidence: 99%

Plant viruses and biomolecular condensates: novel perspectives in virus replication strategies

May

2024

New Phytologist

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Biomolecular condensates are dynamic, membraneless, cellular compartments formed by the reversible assembly of proteins, nucleic acids, and other biomolecules (Hyman et al., 2014). The functions of these condensates in cellular processes have been extensively studied in mammalian systems (Yoshizawa et al., 2020), but similar studies in plants have traditionally lagged far behind. However, in the past c. 5 years, a flurry of research in plants has revealed that biomolecular condensates play crucial roles in cellular organization and functions. These functions are wide-ranging and include flowering, stress responses, RNA processing, and DNA damage, among others (Liu et al., 2024). Like cellular proteins, viral proteins can form condensates during infection, contributing significantly to virus replication cycles. While the functions of biomolecular condensates formed by viral pathogens in mammals are well-documented (Zhang et al., 2023), our understanding of the functions of condensates formed by plant viruses is limited. In an article published in this issue of New Phytologist, Lin & Nagy (2024; 1917-1935 demonstrate that biomolecular condensates formed during Tomato bushy stunt virus (TBSV, Tombusviridae family) infection sequester glycolytic enzymes and increase local adenosine triphosphate (ATP) concentrations for virus replication. These findings advance our understanding of the pro-viral roles of biomolecular condensates, broadening possibilities for how plant viruses concentrate biomolecules beyond traditional membrane-bound virus inclusions formed during infection.

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CASC3 Biomolecular Condensates Restrict Turnip Crinkle Virus by Limiting Host Factor Availability

Cited by 6 publications

References 79 publications

Co-opted cytosolic host proteins form unique condensate substructures within the membranous tombusviral replication organelles

Co-opted cytosolic host proteins form unique condensate substructures within the membranous tombusviral replication organelles

Co‐opted cytosolic proteins form condensate substructures within membranous replication organelles of a positive‐strand RNA virus

Plant viruses and biomolecular condensates: novel perspectives in virus replication strategies

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