The Plant Host Can Affect the Encapsidation of Brome Mosaic Virus (BMV) RNA: BMV Virions Are Surprisingly Heterogeneous

Field-grown wheat (Triticum aestivum L.) plants can be co-infected by multiple viruses, including wheat streak mosaic virus (WSMV), Triticum mosaic virus (TriMV), brome mosaic virus (BMV), and barley stripe mosaic virus (BSMV). These viruses belong to four different genera in three different families and are, hence, genetically divergent. However, the impact of potential co-infections with two, three, or all four of them on the viruses themselves, as well as the wheat host, has yet to be examined. This study examined bi-, tri-, and quadripartite interactions among these viruses in wheat for disease development and accumulation of viral genomic RNAs, in comparison with single virus infections. Co-infection of wheat by BMV and BSMV resulted in BMV-like symptoms with a drastic reduction in BSMV genomic RNA copies and coat protein accumulation, suggesting an antagonism-like effect exerted by BMV toward BSMV. However, co-infection of either BMV or BSMV with WSMV or TriMV led to more severe disease than singly infected wheat, but with a decrease or no significant change in titers of interacting viruses in the presence of BMV or BSMV, respectively. These results were in stark contrast with exacerbated disease phenotype accompanied with enhanced virus titers caused by WSMV and TriMV co-infection. Co-infection of wheat by WSMV, TriMV, and BMV or BSMV resulted in enhanced synergistic disease accompanied by increased accumulation of TriMV and BMV but not WSMV or BSMV. Quadripartite interactions in co-infected wheat by all four viruses resulted in very severe disease synergism, leading to the death of the most infected plants, but paradoxically, a drastic reduction in BSMV titer. Our results indicate that interactions among different viruses infecting the same plant host are more complex than previously thought, do not always entail increases in virus titers, and likely involve multiple mechanisms. These findings lay the foundation for additional mechanistic dissections of synergistic interactions among unrelated plant viruses.

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Identification of Sequences in Brome Mosaic Virus Replicase Protein 1a That Mediate Association with Endoplasmic Reticulum Membranes

Journal of Virology ◽

10.1128/jvi.75.24.12370-12381.2001 ◽

2001 ◽

Vol 75 (24) ◽

pp. 12370-12381 ◽

Cited By ~ 87

Author(s):

Johan A. den Boon ◽

Jianbo Chen ◽

Paul Ahlquist

Keyword(s):

Endoplasmic Reticulum ◽

Mosaic Virus ◽

Fluorescent Protein ◽

Rna Replication ◽

Brome Mosaic Virus ◽

Membrane Association ◽

Plant Host ◽

Multifunctional Protein ◽

Positive Strand Rna ◽

Rna Capping

ABSTRACT RNA replication of all positive-strand RNA viruses is closely associated with intracellular membranes. Brome mosaic virus (BMV) RNA replication occurs on the perinuclear region of the endoplasmic reticulum (ER), both in its natural plant host and in the yeastSaccharomyces cerevisiae. The only viral component in the BMV RNA replication complex that localizes independently to the ER is 1a, a multifunctional protein with an N-terminal RNA capping domain and a C-terminal helicase-like domain. The other viral replication components, the RNA polymerase-like protein 2a and the RNA template, depend on 1a for recruitment to the ER. We show here that, in membrane extracts, 1a is fully susceptible to proteolytic digestion in the absence of detergent and thus, a finding consistent with its roles in RNA replication, is wholly or predominantly on the cytoplasmic face of the ER with no detectable lumenal protrusions. Nevertheless, 1a association with membranes is resistant to high-salt and high-pH treatments that release most peripheral membrane proteins. Membrane flotation gradient analysis of 1a deletion variants and 1a segments fused to green fluorescent protein (GFP) showed that sequences in the N-terminal RNA capping module of 1a mediate membrane association. In particular, a region C-terminal to the core methyltransferase homology was sufficient for high-affinity ER membrane association. Confocal immunofluorescence microscopy showed that even though these determinants mediate ER localization, they fail to localize GFP to the narrow region of the perinuclear ER, where full-length 1a normally resides. Instead, they mediate a more globular or convoluted distribution of ER markers. Thus, additional sequences in 1a that are distinct from the primary membrane association determinants contribute to 1a's normal subcellular distribution, possibly through effects on 1a conformation, orientation, or multimerization on the membrane.

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Virus-Like Particles Produced Using the Brome Mosaic Virus Recombinant Capsid Protein Expressed in a Bacterial System

International Journal of Molecular Sciences ◽

10.3390/ijms22063098 ◽

2021 ◽

Vol 22 (6) ◽

pp. 3098

Author(s):

Aleksander Strugała ◽

Jakub Jagielski ◽

Karol Kamel ◽

Grzegorz Nowaczyk ◽

Marcin Radom ◽

...

Keyword(s):

Ionic Strength ◽

Mosaic Virus ◽

Capsid Protein ◽

Self Assembly ◽

Environmental Changes ◽

Expression System ◽

Brome Mosaic Virus ◽

Virus Like Particles ◽

Assembly Principles ◽

The Impact

Virus-like particles (VLPs), due to their nanoscale dimensions, presence of interior cavities, self-organization abilities and responsiveness to environmental changes, are of interest in the field of nanotechnology. Nevertheless, comprehensive knowledge of VLP self-assembly principles is incomplete. VLP formation is governed by two types of interactions: protein–cargo and protein–protein. These interactions can be modulated by the physicochemical properties of the surroundings. Here, we used brome mosaic virus (BMV) capsid protein produced in an E. coli expression system to study the impact of ionic strength, pH and encapsulated cargo on the assembly of VLPs and their features. We showed that empty VLP assembly strongly depends on pH whereas ionic strength of the buffer plays secondary but significant role. Comparison of VLPs containing tRNA and polystyrene sulfonic acid (PSS) revealed that the structured tRNA profoundly increases VLPs stability. We also designed and produced mutated BMV capsid proteins that formed VLPs showing altered diameters and stability compared to VLPs composed of unmodified proteins. We also observed that VLPs containing unstructured polyelectrolyte (PSS) adopt compact but not necessarily more stable structures. Thus, our methodology of VLP production allows for obtaining different VLP variants and their adjustment to the incorporated cargo.

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Duplex-RT-PCR assay for the simultaneous detection and discrimination of Brome mosaic virus and Cocksfoot mottle virus in cereal plants

Molecular Biology Reports ◽

10.1007/s11033-021-06485-9 ◽

2021 ◽

Author(s):

Katarzyna Trzmiel

Keyword(s):

Mosaic Virus ◽

Simultaneous Detection ◽

Brome Mosaic Virus ◽

Mottle Virus ◽

Grass Species ◽

Replicase Gene ◽

Rt Pcr ◽

Pcr Products ◽

Cereal Plants ◽

Cocksfoot Mottle Virus

AbstractBrome mosaic virus (BMV) and cocksfoot mottle virus (CfMV) are pathogens of grass species including all economically important cereals. Both viruses have been identified in Poland therefore they create a potential risk to cereal crops. In this study, a duplex—reverse transcription—polymerase chain reaction (duplex-RT-PCR) was developed and optimized for simultaneous detection and differentiation of BMV and CfMV as well as for confirmation of their co-infection. Selected primers CfMVdiag-F/CfMVdiag-R and BMV2-F/BMV2-R amplified 390 bp and 798 bp RT-PCR products within coat protein (CP) region of CfMV and replicase gene of BMV, respectively. Duplex-RT-PCR was successfully applied for the detection of CfMV-P1 and different Polish BMV isolates. Moreover, one sample was found to be co-infected with BMV-ML1 and CfMV-ML1 isolates. The specificity of generated RT-PCR products was verified by sequencing. Duplex-RT-PCR, like conventional RT-PCR, was able to detect two viruses occurring in plant tissues in very low concentration (as low as 4.5 pg/µL of total RNA). In contrast to existing methods, newly developed technique offers a significant time and cost-saving advantage. In conclusion, duplex-RT-PCR is a useful tool which can be implemented by phytosanitary services to rapid detection and differentiation of BMV and CfMV.

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Coding Changes in the 3a Cell-to-Cell Movement Gene Can Extend the Host Range of Brome Mosaic Virus Systemic Infection

Virology ◽

10.1006/viro.1995.0057 ◽

1995 ◽

Vol 214 (2) ◽

pp. 464-474 ◽

Cited By ~ 24

Author(s):

WALTER DE JONG ◽

ANTONY CHU ◽

PAUL AHLQUIST

Keyword(s):

Host Range ◽

Mosaic Virus ◽

Cell Movement ◽

Systemic Infection ◽

Brome Mosaic Virus

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CRYO-EM Atomic Model of Brome Mosaic Virus Derived from Direct Electron Detection Images and a Real-Space Model Optimization Protocol

Biophysical Journal ◽

10.1016/j.bpj.2013.11.3321 ◽

2014 ◽

Vol 106 (2) ◽

pp. 600a ◽

Cited By ~ 1

Author(s):

Zhao Wang ◽

Corey Hryc ◽

Benjamin Bammes ◽

Pavel Afonine ◽

Joanita Jakana ◽

...

Keyword(s):

Mosaic Virus ◽

Real Space ◽

Brome Mosaic Virus ◽

Atomic Model ◽

Model Optimization ◽

Space Model ◽

Electron Detection

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cDNA cloning and in vitro transcription of the complete brome mosaic virus genome

Molecular and Cellular Biology ◽

10.1128/mcb.4.12.2876-2882.1984 ◽

1984 ◽

Vol 4 (12) ◽

pp. 2876-2882 ◽

Cited By ~ 1

Author(s):

P Ahlquist ◽

M Janda

Keyword(s):

Mosaic Virus ◽

Transcription Initiation ◽

Direct Sequencing ◽

In Vitro Transcription ◽

Brome Mosaic Virus ◽

Viral Rna ◽

In Vitro Translation ◽

In Vitro Production ◽

Genomic Rnas

Complete cDNA copies of each of the brome mosaic virus genomic RNAs (3.2, 2.8, and 2.1 kilobases in length) were cloned in a novel transcription vector, pPM1, designed to provide exact control of the transcription initiation site. After cleavage at a unique EcoRI site immediately downstream of the inserted cDNA, these clones can be transcribed in vitro by Escherichia coli RNA polymerase to yield complete copies of the brome mosaic virus RNAs. Dideoxy sequencing of 5' transcript cDNA runoff products and direct sequencing of 32P-3'-end-labeled transcripts show that such transcripts initiate at the same 5' position as natural viral RNA and terminate within the EcoRI runoff site after copying the entire viral RNA sequence. When synthesized in the presence of m7GpppG, the transcripts bear the natural capped 5' terminus of brome mosaic virus RNAs. Such transcripts direct the in vitro translation of proteins which coelectrophorese with the translation products of natural brome mosaic virus RNAs. pPM1 should facilitate in vitro production of other viral and nonviral RNAs.

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