Hsc70-2 is required for Beet black scorch virus infection through interaction with replication and capsid proteins

Wang, Xiaoling; Cao, Xiuling; Liu, Min; Zhang, Ruiqi; Zhang, Xin; Gao, Zongyu; Zhao, Xiaofei; Xu, Kai; Li, Dawei; Zhang, Yongliang

doi:10.1038/s41598-018-22778-9

Cited by 21 publications

(19 citation statements)

References 60 publications

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“…The p23 fragments were amplified from infectious TNV‐A C clone pMTC27 (Xi et al, 2008) and cloned into pGDGm at the Bgl II and Sal I sites to generate pGDGm‐p23. The 2a and p23 fragments were cloned into the Bam HI and Spe I sites of pMDC32‐3xFlag (Wang et al, 2018) to generate pMDC32‐2a‐3xFlag and pMDC32‐p23‐3xFlag, respectively. The primers used to construct these plasmids are listed in Table S1.…”

Section: Methodsmentioning

confidence: 99%

Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection

Wang

Jin

et al. 2021

JIPB

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The vacuole is a unique plant organelle that plays an important role in maintaining cellular homeostasis under various environmental stress conditions. However, the effects of biotic stress on vacuole structure has not been examined using three-dimensional (3D) visualization. Here, we performed 3D electron tomography to compare the ultrastructural changes in the vacuole during infection with different viruses. The 3D models revealed that vacuoles are remodeled in cells infected with cucumber mosaic virus (CMV) or tobacco necrosis virus A Chinese isolate (TNV-A C), resulting in the formation of spherules at the periphery of the vacuole. These spherules contain neck-like channels that connect their interior with the cytosol. Confocal microscopy of CMV replication proteins 1a and 2a and TNV-A C auxiliary replication protein p23 showed that all of these proteins localize to the tonoplast. Electron microscopy revealed that the expression of these replication proteins alone is sufficient to induce spherule formation on the tonoplast, suggesting that these proteins play prominent roles in inducing vacuolar membrane remodeling. This is the first report of the 3D structures of viral replication factories built on the tonoplasts. These findings contribute to our understanding of vacuole biogenesis under normal conditions and during assembly of plant (+) RNA virus replication complexes.

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Section: Methodsmentioning

confidence: 99%

Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection

Wang

Jin

et al. 2021

JIPB

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“…The Hsc70-2 protein of N. benthamiana, which is induced upon infection, interacts with the beet black scorch virus (BBSV) replication protein p23. Accordingly, downregulation of Hsc70-2 resulted in reduced BBSV accumulation [ 76 ], likely resulting from reduced virus replication.…”

Section: Host Factors That Determine Virus Susceptibilitymentioning

confidence: 99%

Susceptibility Genes to Plant Viruses

García-Ruíz

2018

Viruses

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Plant viruses use cellular factors and resources to replicate and move. Plants respond to viral infection by several mechanisms, including innate immunity, autophagy, and gene silencing, that viruses must evade or suppress. Thus, the establishment of infection is genetically determined by the availability of host factors necessary for virus replication and movement and by the balance between plant defense and viral suppression of defense responses. Host factors may have antiviral or proviral activities. Proviral factors condition susceptibility to viruses by participating in processes essential to the virus. Here, we review current advances in the identification and characterization of host factors that condition susceptibility to plant viruses. Host factors with proviral activity have been identified for all parts of the virus infection cycle: viral RNA translation, viral replication complex formation, accumulation or activity of virus replication proteins, virus movement, and virion assembly. These factors could be targets of gene editing to engineer resistance to plant viruses.

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“…These approaches, however, are necessarily based on prior discovery acquired by other experimental means. Another experimental strategy successfully used to characterize VRCs has been to pull down tagged viral proteins and analyze the resulting protein populations by mass spectrometry [17-19]. While this last method has provided valid and compelling data, we decided to investigate VRCs from a viral RNA-centered, rather than viral protein-centered, perspective.…”

Section: Introductionmentioning

confidence: 99%

Immunocapture of dsRNA-bound proteins provides insight into tobacco rattle virus replication complexes and reveals Arabidopsis DRB2 to be a wide-spectrum antiviral effector

Incarbone

Clavel

Monsion

et al. 2019

Preprint

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27 Plant RNA viruses form highly organized membrane-bound virus replication complexes 28 (VRCs) to replicate their genome and multiply. VRCs contain both virus-and host-encoded 29 proteins, and have been investigated through a number of experimental approaches, including 30 genetic screens on surrogate model experimental hosts and pull-down of viral replicase 31 proteins. Double-stranded RNA (dsRNA) is an obligate intermediate of the replication 32 process of all RNA viruses and is the trigger of potent antiviral defenses in all eukaryotes. In 33 this work, we describe the use of a GFP-tagged dsRNA-binding protein (B2:GFP) to pull 34 down viral replicating RNA and associated proteins in planta. Following infection of A.35 thaliana constitutively expressing B2:GFP with tobacco rattle virus (TRV), we were able 36 through GFP immunoprecipitation to pull down dsRNA. Mass spectrometry analysis of the 37 dsRNA-B2:GFP-bound proteins from TRV-infected plants revealed the presence of (i) viral 38 proteins such as the replicase, which attested to the successful isolation of VRCs, and (ii) a 39 number of host proteins, some of which have previously been involved in virus infection. To 40 verify whether the identified host proteins localized at TRV replication complexes, we 41 selected nine candidates, generated genetic fusions with RFP, and transiently expressed them 42 in healthy and TRV-infected N. benthamiana stably expressing B2:GFP. Confocal 43 microscopy analysis revealed that eight out of nine candidates showed dramatic re-44 localization upon infection, and seven of these co-localized with B2-labeled TRV replication 45 complexes, providing ample validation for the immunoprecipitation results. We therefore 46 propose B2:GFP-mediated pull down of dsRNA to be a novel and robust method to explore 47 the proteome of VRCs in planta. 48 49 50 51 AUTHOR SUMMARY 52 Viruses are an important class of pathogens that represent a major problem for human, animal 53 and plant health. They hijack the molecular machinery of host cells to complete their 54 replication cycle, a process frequently associated with the production of double-stranded 55 RNA (dsRNA) that is regarded as a universal hallmark of infection by RNA viruses. Here we 56 exploited the capacity of a GFP-tagged dsRNA-binding protein stably expressed in transgenic 57 Arabidopsis to pull down dsRNA and associated proteins upon virus infection. In this manner 58 we specifically captured short and long dsRNA from tobacco rattle virus (TRV) infected 59 plants, and successfully isolated viral proteins such as the replicase, which attested to the 60 successful isolation of virus replication complexes (VRCs). More excitingly, a number of 61 host proteins, some of which have previously been involved in virus infection, were also 62 captured. Remarkably, among a set of nine host candidates that were analyzed, eight showed 63 dramatic re-localization to viral factories upon infection, and seven of these co-localized 64 dsRNA-labeled VRCs. Being compatible with any plant virus as ...

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Hsc70-2 is required for Beet black scorch virus infection through interaction with replication and capsid proteins

Cited by 21 publications

References 60 publications

Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection

Three‐dimensional reconstruction and comparison of vacuolar membranes in response to viral infection

Susceptibility Genes to Plant Viruses

Immunocapture of dsRNA-bound proteins provides insight into tobacco rattle virus replication complexes and reveals Arabidopsis DRB2 to be a wide-spectrum antiviral effector

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