A Non-Canonical Pathway Induced by Externally Applied Virus-Specific dsRNA in Potato Plants.

The external application of double-stranded RNA (dsRNA) has recently been developed as a non-transgenic approach for crop protection against pests and pathogens. This novel and emerging approach has come to prominence due to its safety and environmental benefits. It is generally assumed that the mechanism of dsRNA-mediated antivirus RNA silencing is similar to that of natural RNA interference (RNAi)-based defence against RNA-containing viruses.

The Temporal and Geographical Dynamics of Potato Virus Y Diversity in Russia.

Potato virus Y, an important viral pathogen of potato, has several genetic variants and geographic distributions which could be affected by environmental factors, aphid vectors, and reservoir plants. PVY is transmitted to virus-free potato plants by aphids and passed on to the next vegetative generations through tubers, but the effects of tuber transmission in PVY is largely unknown. By using high-throughput sequencing, we investigated PVY populations transmitted to potato plants by aphids in different climate zones of Russia, namely the Moscow and Astrakhan regions.

Cajal bodies: Evolutionarily conserved nuclear biomolecular condensates with properties unique to plants.

Proper orchestration of the thousands of biochemical processes that are essential to the life of every cell requires highly organized cellular compartmentalization of dedicated microenvironments. There are 2 ways to create this intracellular segregation to optimize cellular function. One way is to create specific organelles, enclosed spaces bounded by lipid membranes that regulate macromolecular flux in and out of the compartment.

Psoroptes ovis-Early Immunoreactive Protein (Pso-EIP-1) a novel diagnostic antigen for sheep scab.

Aims: Serodiagnosis of sheep scab is an established diagnostic method and has become popular in recent years. However, the current diagnostic antigen, Pso o 2, has shown promise as a component of a recombinant vaccine for scab, making it incompatible with discriminating between infected and vaccinated animals (DIVA). Here, we describe the discovery and characterization of a novel Psoroptes ovis immunodiagnostic antigen, P.

CRISPR Applications in Plant Virology: Virus Resistance and Beyond.

Clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated genes (Cas) is a prokaryotic adaptive immune system which has been reprogrammed into a precise, simple, and efficient gene targeting technology. This emerging technology is revolutionizing various areas of life sciences, medicine, and biotechnology and has raised significant interest among plant biologists, both in basic science and in plant protection and breeding. In this review, we describe the basic principles of CRISPR/Cas systems, and how they can be deployed to model plants and crops for the control, monitoring, and study of the mechanistic aspects of plant virus infections.

Interaction of a plant virus protein with the signature Cajal body protein coilin facilitates salicylic acid-mediated plant defence responses.

In addition to well-known roles in RNA metabolism, the nucleolus and Cajal bodies (CBs), both located within the nucleus, are involved in plant responses to biotic and abiotic stress. Previously we showed that plants in which expression of the CB protein coilin is downregulated are more susceptible to certain viruses including tobacco rattle virus (TRV), suggesting a role of coilin in antiviral defence. Experiments with coilin-deficient plants and the deletion mutant of the TRV 16K protein showed that both 16K and coilin are required for restriction of systemic TRV infection.

Virus-Like Particle Facilitated Deposition of Hydroxyapatite Bone Mineral on Nanocellulose after Exposure to Phosphate and Calcium Precursors.

We produced and isolated tobacco mosaic virus-like particles (TMV VLPs) from bacteria, which are devoid of infectious genomes, and found that they have a net negative charge and can bind calcium ions. Moreover, we showed that the TMV VLPs could associate strongly with nanocellulose slurry after a simple mixing step. We sequentially exposed nanocellulose alone or slurries mixed with the TMV VLPs to calcium and phosphate salts and utilized physicochemical approaches to demonstrate that bone mineral (hydroxyapatite) was deposited only in nanocellulose mixed with the TMV VLPs.

Propagation and Isolation of Tobacco Mosaic Virus That Surface Displays Metal Binding and Reducing Peptides for Generation of Gold Nanoparticles.

In this chapter we describe an approach for propagating, isolating and characterizing tobacco mosaic virus (TMV) genetically modified to surface display a metal binding and reducing peptide, and its utilization for the production of free gold metal nanoparticles from metal salt precursors.

Data on a delivery of biomolecules into leaves using different nanoparticles.

Nanoparticles (NPs) have a number of unique properties associated with their ultrasmall size and exhibit many advantages compared with existing plant biotechnology platforms for delivery of proteins, RNA and DNA of various sizes into the plant cells (Arruda et al., 2015; Silva et al., 2010; Martin-Ortigosa et al.

Phytaspase-mediated precursor processing and maturation of the wound hormone systemin.

Peptide hormones are implicated in many important aspects of plant life and are usually synthesized as precursor proteins. In contrast to animals, data for plant peptide hormone maturation are scarce and the specificity of processing enzyme(s) is largely unknown. Here we tested a hypothesis that processing of prosystemin, a precursor of tomato (Solanum lycopersicum) wound hormone systemin, is performed by phytaspases, aspartate-specific proteases of the subtilase family.

Cajal bodies and their role in plant stress and disease responses.

Cajal bodies (CBs) are distinct sub-nuclear structures that are present in eukaryotic living cells and are often associated with the nucleolus. CBs play important roles in RNA metabolism and formation of RNPs involved in transcription, splicing, ribosome biogenesis, and telomere maintenance. Besides these primary roles, CBs appear to be involved in additional functions that may not be directly related to RNA metabolism and RNP biogenesis.

A Genetically Modified Tobacco Mosaic Virus that can Produce Gold Nanoparticles from a Metal Salt Precursor.

We genetically modified tobacco mosaic virus (TMV) to surface display a characterized peptide with potent metal ion binding and reducing capacity (MBP TMV), and demonstrate that unlike wild type TMV, this construct can lead to the formation of discrete 10-40 nm gold nanoparticles when mixed with 3 mM potassium tetrachloroaurate. Using a variety of analytical physicochemical approaches it was found that these nanoparticles were crystalline in nature and stable. Given that the MBP TMV can produce metal nanomaterials in the absence of chemical reductants, it may have utility in the green production of metal nanomaterials.

Biosynthesis of stable iron oxide nanoparticles in aqueous extracts of Hordeum vulgare and Rumex acetosa plants.

We report the synthesis and characterization of amorphous iron oxide nanoparticles from iron salts in aqueous extracts of monocotyledonous (Hordeum vulgare) and dicotyledonous (Rumex acetosa) plants. The nanoparticles were characterized by TEM, absorbance spectroscopy, SAED, EELS, XPS, and DLS methods and were shown to contain mainly iron oxide and iron oxohydroxide. H.

Coilin, the signature protein of Cajal bodies, differentially modulates the interactions of plants with viruses in widely different taxa.

Cajal bodies (CBs) are distinct nuclear bodies physically and functionally associated with the nucleolus. In addition to their traditional function in coordinating maturation of certain nuclear RNAs, CBs participate in cell cycle regulation, development, and regulation of stress responses. A key "signature" component of CBs is coilin, the scaffolding protein essential for CB formation and function.

The use of tobacco mosaic virus and cowpea mosaic virus for the production of novel metal nanomaterials.

Due to the nanoscale size and the strictly controlled and consistent morphologies of viruses, there has been a recent interest in utilizing them in nanotechnology. The structure, surface chemistries and physical properties of many viruses have been well elucidated, which have allowed identification of regions of their capsids which can be modified either chemically or genetically for nanotechnological uses. In this review we focus on the use of such modifications for the functionalization and production of viruses and empty viral capsids that can be readily decorated with metals in a highly tuned manner.

Dynamic localization of two tobamovirus ORF6 proteins involves distinct organellar compartments.

ORF6 is a small gene that overlaps the movement and coat protein genes of subgroup 1a tobamoviruses. The ORF6 protein of tomato mosaic virus (ToMV) strain L (L-ORF6), interacts in vitro with eukaryotic elongation factor 1α, and mutation of the ORF6 gene of tobacco mosaic virus (TMV) strain U1 (U1-ORF6) reduces the pathogenicity in vivo of TMV, whereas expression of this gene from two other viruses, tobacco rattle virus (TRV) and potato virus X (PVX), increases their pathogenicity. In this work, the in vivo properties of the L-ORF6 and U1-ORF6 proteins were compared to identify sequences that direct the proteins to different subcellular locations and also influence virus pathogenicity.

The extreme N-terminal domain of a hordeivirus TGB1 movement protein mediates its localization to the nucleolus and interaction with fibrillarin.

The hordeiviral movement protein encoded by the first gene of the triple gene block (TGBp1) of Poa semilatent virus (PSLV), interacts with viral genomic RNAs to form RNP particles which are considered to be a form of viral genome capable of cell-to-cell and long-distance transport in infected plants. The PSLV TGBp1 contains a C-terminal NTPase/helicase domain (HELD) and an N-terminal extension region consisting of two structurally and functionally distinct domains: an extreme N-terminal domain (NTD) and an internal domain (ID). This study demonstrates that transient expression of TGBp1 fused to GFP in Nicotiana benthamiana leaves results in faint but obvious fluorescence in the nucleolus in addition to cytosolic distribution.

Domain organization of the N-terminal portion of hordeivirus movement protein TGBp1.

Three 'triple gene block' proteins known as TGBp1, TGBp2 and TGBp3 are required for cell-to-cell movement of plant viruses belonging to a number of genera including Hordeivirus. Hordeiviral TGBp1 interacts with viral genomic RNAs to form ribonucleoprotein (RNP) complexes competent for translocation between cells through plasmodesmata and over long distances via the phloem. Binding of hordeivirus TGBp1 to RNA involves two protein regions, the C-terminal NTPase/helicase domain and the N-terminal extension region.

Cylindrical inclusion protein of potato virus A is associated with a subpopulation of particles isolated from infected plants.

Potato virus A (PVA) particles were purified by centrifugation through a 30 % sucrose cushion and the pellet (P1) was resuspended and sedimented through a 5-40 % sucrose gradient. The gradient separation resulted in two different virus particle populations: a virus fraction (F) that formed a band in the gradient and one that formed a pellet (P2) at the bottom of the gradient. All three preparations contained infectious particles that retained their integrity when visualized by electron microscopy (EM).

Caspase-resistant VirD2 protein provides enhanced gene delivery and expression in plants.

Agrobacterium tumefaciens VirD2 protein is one of the key elements of Agrobacterium-mediated plant transformation, a process of transfer of T-DNA sequence from the Agrobacterium tumour inducing plasmid into the nucleus of infected plant cells and its integration into the host genome. The VirD2 protein has been shown to be a substrate for a plant caspase-like protease activity (PCLP) in tobacco. We demonstrate here that mutagenesis of the VirD2 protein to prevent cleavage by PCLP increases the efficiency of reporter gene transfer and expression.

An unusual structure at one end of potato potyvirus particles.

The particles of potato virus Y (PVY) and potato virus A (PVA) potyviruses are helically constructed filaments that are thought to contain a single type of coat protein subunit. Examination of negatively stained virions by electron microscopy reveals flexuous rod-shaped particles with no obvious terminal structures. It is known that some helically constructed rod-shaped virus particles incorporate additional minor protein components, which form stable complexes that mediate particle disassembly, movement or transmission by vectors.

Molecular interactions between a plant virus movement protein and RNA: force spectroscopy investigation.

RNA-protein interactions are fundamental for different aspects of molecular biology such as gene expression, assembly of biomolecular complexes or macromolecular transport. The 3a movement protein (MP) of a plant virus, Cucumber mosaic virus (CMV), forms ribonucleoprotein (RNP) complexes with viral RNA, capable of trafficking from cell-to-cell throughout the infected plant only in the presence of the CMV capsid protein (CP). However, deletion of the C-terminal 33 amino acid residues of the CMV MP (in the mutant designated 3aDeltaC33 MP) resulted in CP-independent cell-to-cell movement.

Complex molecular architecture of beet yellows virus particles.

Closteroviruses possess exceptionally long filamentous virus particles that mediate protection and active transport of the genomic RNA within infected plants. These virions are composed of a long "body" and short "tail" whose principal components are the major and minor capsid proteins, respectively. Here we use biochemical, genetic, and ultrastructural analyses to dissect the molecular composition and architecture of particles of beet yellows virus, a closterovirus.

The C-terminal 33 amino acids of the cucumber mosaic virus 3a protein affect virus movement, RNA binding and inhibition of infection and translation.

The capsid protein (CP) of Cucumber mosaic virus (CMV) is required for cell-to-cell movement, mediated by the 3a movement protein (MP). Deletion of the C-terminal 33 amino acids of the CMV 3a MP (in the mutant designated 3aDeltaC33 MP) resulted in CP-independent cell-to-cell movement, but not long-distance movement. RNA-binding studies done in vitro using isolated bacterially expressed MP showed that the 3aDeltaC33 MP bound RNA more strongly, with fewer regions sensitive to RNase and formed cooperatively bound complexes at lower ratios of protein : RNA than the wild-type (wt) 3a MP.

A plant caspase-like protease activated during the hypersensitive response.

To test the hypothesis that caspase-like proteases exist and are critically involved in the implementation of programmed cell death (PCD) in plants, a search was undertaken for plant caspases activated during the N gene-mediated hypersensitive response (HR; a form of pathogen-induced PCD in plants) in tobacco plants infected with Tobacco mosaic virus (TMV). For detection, characterization, and partial purification of a tobacco caspase, the Agrobacterium tumefaciens VirD2 protein, shown here to be cleaved specifically at two sites (TATD and GEQD) by human caspase-3, was used as a target. In tobacco leaves, specific proteolytic processing of the ectopically produced VirD2 derivatives at these sites was found to occur early in the course of the HR triggered by TMV.