Nanocarriers are widely used for efficient delivery of different cargo into mammalian cells; however, delivery into plant cells remains a challenging issue due to physical and mechanical barriers such as the cuticle and cell wall. Here, we discuss recent progress on biodegradable and biosafe nanomaterials that were demonstrated to be applicable to the delivery of nucleic acids into plant cells. This review covers studies the object of which is the plant cell and the cargo for the nanocarrier is either DNA or RNA. The following nanoplatforms that could be potentially used for nucleic acid foliar delivery via spraying are discussed: mesoporous silica nanoparticles, layered double hydroxides (nanoclay), carbon-based materials (carbon dots and single-walled nanotubes), chitosan and, finally, cell-penetrating peptides (CPPs). Hybrid nanomaterials, for example, chitosan- or CPP-functionalized carbon nanotubes, are taken into account. The selected nanocarriers are analyzed according to the following aspects: biosafety, adjustability for the particular cargo and task (e.g., organelle targeting), penetration efficiency and ability to protect nucleic acid from environmental and cellular factors (pH, UV, nucleases, etc.) and to mediate the gradual and timely release of cargo. In addition, we discuss the method of application, experimental system and approaches that are used to assess the efficiency of the tested formulation in the overviewed studies. This review presents recent progress in developing the most promising nanoparticle-based materials that are applicable to both laboratory experiments and field applications.

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. We analyzed sprouts from the tubers produced by field-infected plants to investigate the impact of tuber transmission on PVY genetics. We found a significantly higher diversity of PVY isolates in the Astrakhan region, where winters are shorter and milder and summers are warmer compared to the Moscow region. While five PVY types, NTNa, NTNb, N:O, N-Wi, and SYR-I, were present in both regions, SYRI-II, SYRI-III, and 261-4 were only found in the Astrakhan region. All these recombinants were composed of the genome sections derived from PVY types O and N, but no full-length sequences of such types were present. The composition of the PVY variants in the tuber sprouts was not always the same as in their parental plants, suggesting that tuber transmission impacts PVY genetics.

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. There is, however, no direct evidence to support this idea. Here, we provide data on the high-throughput sequencing (HTS) analysis of small non-coding RNAs (sRNA) as hallmarks of RNAi induced by infection with the RNA-containing potato virus Y (PVY) and also by exogenous application of dsRNA which corresponds to a fragment of the PVY genome. Intriguingly, in contrast to PVY-induced production of discrete 21 and 22 nt sRNA species, the externally administered PVY dsRNA fragment led to generation of a non-canonical pool of sRNAs, which were present as ladders of ~18–30 nt in length; suggestive of an unexpected sRNA biogenesis pathway. Interestingly, these non-canonical sRNAs are unable to move systemically and also do not induce transitive amplification. These findings may have significant implications for further developments in dsRNA-mediated crop protection.

Nanoparticles could improve the bioavailability of active agents of various natures to human, animal, and plant tissues. In this work, we compared two methods on the synthesis of calcium phosphate nanoparticles (CaPs), differed by the synthesis temperature, pH, and concentration of the stabilizing agent, and explored the possibilities of incorporation of a low-molecular-weight peptide analogue enalaprilat, the enzyme superoxide dismutase 1 (SOD1), as well as DNA and dsRNA into these particles, by coprecipitation and sorption. CaPs obtained with and without cooling demonstrated the highest inclusion efficiency for enalaprilat upon coprecipitation: 250 ± 10 μg/mg of CaPs and 340 ± 30 μg/mg of CaPs, respectively. Enalaprilat sorption on the preliminarily formed CaPs was much less effective. SOD1 was only able to coprecipitate with CaPs upon cooling, with SOD1 loading 6.6 ± 2 μg/mg of CaPs. For the incorporation of DNA, the superiority of the sorption method was demonstrated, allowing loading of up to 88 μg/mg of CaPs. The ability of CaPs to incorporate dsRNa by sorption was also demonstrated by electrophoresis and atomic force microscopy. These results could have important implications for the development of the roots for incorporating substances of different natures into CaPs for agricultural and medical applications.

ADP-ribosylation (ADPRylation) is a mechanism which post-translationally modifies proteins in eukaryotes in order to regulate a broad range of biological processes including programmed cell death, cell signaling, DNA repair, and responses to biotic and abiotic stresses. Poly(ADP-ribosyl) polymerases (PARPs) play a key role in the process of ADPRylation, which modifies target proteins by attaching ADP-ribose molecules. Here, we investigated whether and how PARP1 and PARylation modulate responses of Nicotiana benthamiana plants to methyl viologen (MV)-induced oxidative stress. It was found that the burst of reactive oxygen species (ROS), cell death, and loss of tissue viability invoked by MV in N. benthamiana leaves was significantly delayed by both the RNA silencing of the PARP1 gene and by applying the pharmacological inhibitor 3-aminobenzamide (3AB) to inhibit PARylation activity. This in turn reduced the accumulation of PARylated proteins and significantly increased the gene expression of major ROS scavenging enzymes including SOD (NbMnSOD; mitochondrial manganese SOD), CAT (NbCAT2), GR (NbGR), and APX (NbAPX5), and inhibited cell death. This mechanism may be part of a broader network that regulates plant sensitivity to oxidative stress through various genetically programmed pathways.

Mass spectrometry-based proteome analysis usually implies matching mass spectra of proteolytic peptides to amino acid sequences predicted from nucleic acid sequences. At the same time, due to the stochastic nature of the method when it comes to proteome-wide analysis, in which only a fraction of peptides are selected for sequencing, the completeness of protein sequence identification is undermined. Likewise, the reliability of peptide variant identification in proteogenomic studies is suffering. We propose a way to interpret shotgun proteomics results, specifically in data-dependent acquisition mode, as protein sequence coverage by multiple reads, just as it is done in the field of nucleic acid sequencing for the calling of single nucleotide variants. Multiple reads for each position in a sequence could be provided by overlapping distinct peptides, thus, confirming the presence of certain amino acid residues in the overlapping stretch with much lower false discovery rate than conventional 1%. The source of overlapping distinct peptides are, first, miscleaved tryptic peptides in combination with their properly cleaved counterparts, and, second, peptides generated by several proteases with different specificities after the same specimen is subject to parallel digestion and analyzed separately. We illustrate this approach using publicly available multiprotease proteomic datasets and our own data generated for HEK-293 cell line digests obtained using trypsin, LysC and GluC proteases. From 5000 to 8000 protein groups are identified for each digest corresponding to up to 30% of the whole proteome coverage. Most of this coverage was provided by a single read, while up to 7% of the observed protein sequences were covered two-fold and more. The proteogenomic analysis of HEK-293 cell line revealed 36 peptide variants associated with SNP, seven of which were supported by multiple reads. The efficiency of the multiple reads approach depends strongly on the depth of proteome analysis, the digesting features such as the level of miscleavages, and will increase with the number of different proteases used in parallel proteome digestion. Graphical