Insights into the genetic variability and evolutionary dynamics of tomato spotted wilt orthotospovirus in China.

BACKGROUND: Viral diseases are posing threat to annual production and quality of tobacco in China. Recently, tomato spotted wilt orthotospovirus (TSWV) has been reported to infect three major crops including tobacco. Current study was aimed to investigate the population dynamics and molecular diversity of the TSWV. In the current study, to assess and identify the prevalence and evolutionary history of TSWV in tobacco crops in China, full-length genome sequences of TSWV isolates from tobacco, were identified and analyzed. METHODS: After trimming and validation, sequences of new isolates were submitted to GenBank. We identified the full-length genomes of ten TSWV isolates, infecting tobacco plants from various regions of China. Besides these, six isolates were partially sequenced. Phylogenetic analysis was performed to assess the relativeness of newly identified sequences and corresponding sequences from GenBank. Recombination and population dynamics analysis was performed using RDP4, RAT, and statistical estimation. Reassortment analysis was performed using MegaX software. RESULTS: Phylogenetic analysis of 41 newly identified sequences, depicted that the majority of the Chinese isolates have separate placement in the tree. RDP4 software predicted that RNA M of newly reported isolate YNKM-2 had a recombinant region spanning from 3111 to 3811 bp. The indication of parental sequences (YNKMXD and YNHHKY) from newly identified isolates, revealed the conservation of local TSWV population. Genetic diversity and population dynamics analysis also support the same trend. RNA M was highlighted to be more capable of mutating or evolving as revealed by data obtained from RDP4, RAT, population dynamics, and phylogenetic analyses. Reassortment analysis revealed that it might have happened in L segment of TSWV isolate YNKMXD (reported herein). CONCLUSION: Taken together, this is the first detailed study revealing the pattern of TWSV genetic diversity, and population dynamics helping to better understand the ability of this pathogen to drastically reduce the tobacco production in China. Also, this is a valuable addition to the existing worldwide profile of TSWV, especially in China, where a few studies related to TSWV have been reported including only one complete genome of this virus isolated from tobacco plants.

Non-transgenic, PAMAM co-delivery DNA of interactive proteins NbCRVP and NbCalB endows Nicotiana benthamiana with a stronger antiviral effect to RNA viruses.

BACKGROUND: Viral diseases continue to pose a major threat to the world's commercial crops. The in-depth exploration and efficient utilization of resistance proteins have become crucial strategies for their control. However, current delivery methods for introducing foreign DNA suffer from host range limitations, low transformation efficiencies, tissue damage, or unavoidable DNA integration into the host genome. The nanocarriers provides a convenient channel for the DNA delivery and functional utilization of disease-resistant proteins. RESULTS: In this research, we identified a cysteine-rich venom protein (NbCRVP) in Nicotiana benthamiana for the first time. Virus-induced gene silencing and transient overexpression clarified that NbCRVP could inhibit the infection of tobacco mosaic virus, potato virus Y, and cucumber mosaic virus, making it a broad-spectrum antiviral protein. Yeast two-hybrid assay, co-immunoprecipitation, and bimolecular fluorescence complementation revealed that calcium-dependent lipid-binding (CaLB domain) family protein (NbCalB) interacted with NbCRVP to assist NbCRVP playing a stronger antiviral effect. Here, we demonstrated for the first time the efficient co-delivery of DNA expressing NbCRVP and NbCalB into plants using poly(amidoamine) (PAMAM) nanocarriers, achieving stronger broad-spectrum antiviral effects. CONCLUSIONS: Our work presents a tool for species-independent transfer of two interacting protein DNA into plant cells in a specific ratio for enhanced antiviral effect without transgenic integration, which further demonstrated new strategies for nanocarrier-mediated DNA delivery of disease-resistant proteins.

In Vivo Production of dsRNA Using Bacteriophage ϕ6 in Pseudomonas syringae Cit7 Cells.

RNA interference (RNAi), also known as post-transcriptional gene silencing (PTGS), is one of the emerging genetic engineering techniques to effectively silence or inhibit the expression of target genes. This chapter describes a method for in vivo production of dsRNA in non-pathogenic Pseudomonas syringae strains using phage ϕ6 RNA-dependent RNA polymerase, extraction and purification of dsRNA from bacterial solution, and the use of dsRNA to induce silencing of green fluorescent protein (GFP) in transgenic Nicotiana benthamiana.

Nanoparticle-dsRNA Treatment of Pollen and Root Systems of Diseased Plants Effectively Reduces the Rate of Tobacco Mosaic Virus in Contemporary Seeds.

Most crop viruses are carried and spread by seeds. Virus-infected seeds are seed-borne viral disease infections, and thus, reducing the rate of seed infection is an urgent problem in the seed-production industry. The objective of this study was to use nanoparticles (NPs) to directly deliver dsRNA into plants or pollen to initiate RNA interference (RNAi) to reduce viral carryover in seeds. Chitosan quaternary ammonium salt (HACC), complexed with dsRNAs, was selected for targeting the genes for the tobacco mosaic virus (TMV) coat protein (CP) and TMV RNA-dependent RNA polymerase (RdRP) to form HACC-dsRNA NPs. These NP-based dsRNAs were delivered to the plants using four different methods, including infiltration, spraying, root soaking, and pollen internalization. All four methods were able to reduce the seed-carrying rate of offspring seeds of the TMV-infected plants, with pollen internalization being the most effective in reducing the TMV-carrying rate from 95.1 to 61.1% in the control group. By measuring the plant uptake of fluorescence-labeled NPs and dsRNAs, the transportation of the HACC-dsRNA NPs into the plants was observed, and the uptake of dsRNA in combination with small RNA sequencing was further confirmed, resulting in the silencing of homologous RNA molecules during the topical application. The results demonstrated that the incidence of TMV infection was reduced by various degrees via RNAi induction without the need to develop transgenic plants. These results demonstrate the advantages of NP-based RNAi technology in breeding for disease resistance and developing a new strategy for virus-resistant breeding in plants.

Evaluation of the anti-viral efficacy of three different dsRNA nanoparticles against potato virus Y using various delivery methods.

Nanoparticles (NPs) derived from RNA interference (RNAi) are considered a potentially revolutionary technique in the field of plant protection in the future. However, the application of NPs in RNAi is hindered by the conflict between the high cost of RNA production and the large quantity of materials required for field application. This study aimed to evaluate the antiviral efficacy of commercially available nanomaterials, such as chitosan quaternary ammonium salt (CQAS), amine functionalized silica nano powder (ASNP), and carbon quantum dots (CQD), that carried double-stranded RNA (dsRNA) via various delivery methods, including infiltration, spraying, and root soaking. ASNP-dsRNA NPs are recommended for root soaking, which is considered the most effective method of antiviral compound application. The most effective antiviral compound tested was CQAS-dsRNA NPs delivered by root soaking. Using fluorescence, FITC-CQAS-dsCP-Cy3, and CQD-dsCP-Cy3 NPs demonstrated the uptake and transport pathways of dsRNA NPs in plants when applied to plants in different modes. The duration of protection with NPs applied in various modes was then compared, providing references for evaluating the retention period of various types of NPs. All three types of NPs effectively silenced genes in plants and afforded at least 14 days of protection against viral infection. Particularly, CQD-dsRNA NPs could protect systemic leaves for 21 days following spraying.

Analysis of lysine acetylation in tomato spot wilt virus infection in Nicotiana benthamiana.

Introduction: Kac is a model for all acylation modification studies. Kac plays a critical role in eukaryotes and prokaryotes. It is mainly involved in six major biological functions: gene expression, signal transduction, cell development, protein conversion, metabolism, and metabolite transport. Method: We investigated and compared the acetylation modification of proteins in healthy and tomato spot wilt virus (TSWV)-infected Nicotiana benthamiana leaves. Result: We identified 3,418 acetylated lysine sites on 1962 proteins acetylation of proteins in the TSWV-infected and control groups were compared; it was observed that 408 sites on 294 proteins were upregulated and 284 sites on 219 proteins (involved in pentose phosphate, photosynthesis, and carbon fixation in photosynthesis) were downregulated after the infection. Overall, 35 conserved motifs were identified, of which xxxkxxxxx_K_ Rxxxxxxxxx represented 1,334 (31.63%) enrichment motifs and was the most common combination. Bioinformatic analysis revealed that most of the proteins with Kac sites were located in the chloroplast and cytoplasm. They were involved in biological processes, such as cellular and metabolic processes. Discussion: In conclusion, our results revealed that Kac may participate in the regulation of TSWV infection in N. benthamiana.

NbMLP43 Ubiquitination and Proteasomal Degradation via the Light Responsive Factor NbBBX24 to Promote Viral Infection.

The ubiquitin-proteasome system (UPS) plays an important role in virus-host interactions. However, the mechanism by which the UPS is involved in innate immunity remains unclear. In this study, we identified a novel major latex protein-like protein 43 (NbMLP43) that conferred resistance to Nicotiana benthamiana against potato virus Y (PVY) infection. PVY infection strongly induced NbMLP43 transcription but decreased NbMLP43 at the protein level. We verified that B-box zinc finger protein 24 (NbBBX24) interacted directly with NbMLP43 and that NbBBX24, a light responsive factor, acted as an essential intermediate component targeting NbMLP43 for its ubiquitination and degradation via the UPS. PVY, tobacco mosaic virus, (TMV) and cucumber mosaic virus (CMV) infections could promote NbMLP43 ubiquitination and proteasomal degradation to enhance viral infection. Ubiquitination occurred at lysine 38 (K38) within NbMLP43, and non-ubiquitinated NbMLP43(K38R) conferred stronger resistance to RNA viruses. Overall, our results indicate that the novel NbMLP43 protein is a target of the UPS in the competition between defense and viral anti-defense and enriches existing theoretical studies on the use of UPS by viruses to promote infection.

Genome-wide analysis of MYB family in Nicotiana benthamiana and the functional role of the key members in resistance to Bemisia tabaci.

MYB transcription factors (TFs) play a key role in plant resistance to abiotic and biotical stresses. However, little is currently known about their involvement in the plant defense to piercing-sucking insects. Here, we studied the MYB TFs that responded to and resisted Bemisia tabaci whitefly in the model plant Nicotiana benthamiana. Firstly, a total of 453 NbMYB TFs in N. benthamiana genome were identified and 182 R2R3-MYB TFs were analyzed for molecular characteristics, phylogenetic analysis, genetic structure, motif composition, and cis-elements. Then, six stress-related NbMYB genes were selected for further study. The expression pattern shows they were highly expressed in mature leaves and intensively induced upon whitefly attack. Combined with bioinformatic analysis, overexpression, ß-Glucuronidase (GUS) assay, and virus-induced silencing tests, we determined the transcriptional regulation of these NbMYBs on the genes in lignin biosynthesis and SA-signaling pathways. Meanwhile, we tested the performance of whitefly on plants with increased or silenced NbMYB genes expression and found that NbMYB42, NbMYB107, NbMYB163, and NbMYB423 were resistant to whitefly. Our results contribute to a comprehensive understanding of the MYB TFs in N. benthamiana. Furthermore, our findings will facilitate further studies on the role of MYB TFs in the interaction between plants and piercing-sucking insects.

Yiyi Fuzi Baijiang Powder Alleviates Dextran Sulfate Sodium-Induced Ulcerative Colitis in Rats via Inhibiting the TLR4/NF-κB/NLRP3 Inflammasome Signaling Pathway to Repair the Intestinal Epithelial Barrier, and Modulating Intestinal Microbiota.

Ulcerative colitis (UC) is a chronic non-specific inflammatory disease of the intestine, which is prone to recurrence and difficult to cure. Yiyi Fuzi Baijiang powder (YFBP), as a classic Chinese herbal formula, is commonly used in the clinical treatment of UC. However, its potential mechanism remains unclear. In this study, we investigated the mechanism by which YFBP exerts a therapeutic effect against UC. Firstly, we used network pharmacology to screen the active ingredients and potential targets of YFBP and constructed a "drug-ingredient-target" network. Based on bioinformatics, we searched for differentially expressed genes (DEGs) associated with UC and obtained common targets. The core targets of YFBP in the treatment of UC were identified using a protein-protein interaction (PPI) network, and molecular docking techniques were used to evaluate the binding energies of the core targets and corresponding ingredients. Enrichment analysis by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) revealed that YFBP exerted therapeutic effects by regulating multiple inflammatory pathways including TLR4, NF-κB, and TNF. Secondly, an experimental study was carried out in vivo for verification. Our results demonstrated that YFBP could effectively improve the symptoms and intestinal pathological of UC rats. Further study showed that YFBP could significantly downregulate the expressions of TLR4 and p-NF-κB p65 in UC rats, inhibit the activation of NLRP3 inflammasome, reduce the levels of IL-1ß and TNF-α, and then upregulate the expressions of tight junction proteins in intestinal epithelial cells. In addition, YFBP could improve the intestinal microbial community. In conclusion, our study revealed that YFBP had a good therapeutic effect on UC, and its mechanism might be related to the inhibition of the TLR4/NF-κB/NLRP3 inflammasome signaling pathway to repair intestinal epithelial barrier and the modulation of intestinal microbiota.

Unraveling the regulatory network of miRNA expression in Potato Y virus-infected of Nicotiana benthamiana using integrated small RNA and transcriptome sequencing.

Potato virus Y (PVY) disease is a global problem that causes significant damage to crop quality and yield. As traditional chemical control methods are ineffective against PVY, it is crucial to explore new control strategies. MicroRNAs (miRNAs) play a crucial role in plant and animal defense responses to biotic and abiotic stresses. These endogenous miRNAs act as a link between antiviral gene pathways and host immunity. Several miRNAs target plant immune genes and are involved in the virus infection process. In this study, we conducted small RNA sequencing and transcriptome sequencing on healthy and PVY-infected N. benthamiana tissues (roots, stems, and leaves). Through bioinformatics analysis, we predicted potential targets of differentially expressed miRNAs using the N. benthamiana reference genome and the PVY genome. We then compared the identified differentially expressed mRNAs with the predicted target genes to uncover the complex relationships between miRNAs and their targets. This study successfully constructed a miRNA-mRNA network through the joint analysis of Small RNA sequencing and transcriptome sequencing, which unveiled potential miRNA targets and identified potential binding sites of miRNAs on the PVY genome. This miRNA-mRNA regulatory network suggests the involvement of miRNAs in the virus infection process.

NtbHLH49, a jasmonate-regulated transcription factor, negatively regulates tobacco responses to Phytophthora nicotianae.

Tobacco black shank caused by Phytophthora nicotianae is a devastating disease that causes huge losses to tobacco production across the world. Investigating the regulatory mechanism of tobacco resistance to P. nicotianae is of great importance for tobacco resistance breeding. The jasmonate (JA) signaling pathway plays a pivotal role in modulating plant pathogen resistance, but the mechanism underlying JA-mediated tobacco resistance to P. nicotianae remains largely unclear. This work explored the P. nicotianae responses of common tobacco cultivar TN90 using plants with RNAi-mediated silencing of NtCOI1 (encoding the perception protein of JA signal), and identified genes involved in this process by comparative transcriptome analyses. Interestingly, the majority of the differentially expressed bHLH transcription factor genes, whose homologs are correlated with JA-signaling, encode AtBPE-like regulators and were up-regulated in NtCOI1-RI plants, implying a negative role in regulating tobacco response to P. nicotianae. A subsequent study on NtbHLH49, a member of this group, showed that it's negatively regulated by JA treatment or P. nicotianae infection, and its protein was localized to the nucleus. Furthermore, overexpression of NtbHLH49 decreased tobacco resistance to P. nicotianae, while knockdown of its expression increased the resistance. Manipulation of NtbHLH49 expression also altered the expression of a set of pathogen resistance genes. This study identified a set of genes correlated with JA-mediated tobacco response to P. nicotianae, and revealed the function of AtBPE-like regulator NtbHLH49 in regulating tobacco resistance to this pathogen, providing insights into the JA-mediated tobacco responses to P. nicotianae.

Manipulation of Whitefly Behavior by Plant Viruses.

Whiteflies of the Bemisia tabaci complex transmit hundreds of plant viruses belonging to the genera Begomovirus and Crinivirus, among others. Tripartite interactions of whitefly-virus-plant frequently occur during virus infection and transmission. Specifically, virus transmission-related behavior of whitefly, such as preference and feeding, may be altered by viruses and thus exert significant impacts on the outcome of virus spread and epidemics. Here, we provide an overview on the current understanding of the manipulation of whitefly behavior by plant viruses. Plant viruses can significantly modulate whitefly preference and feeding behavior, either directly or in a plant-mediated manner. In general, non-viruliferous whiteflies tend to prefer virus-infected plants, and viruliferous whiteflies are more likely to prefer uninfected plants. In most cases, virus infection of plants and/or whitefly seems to exhibit positive or no effects on whitefly feeding on plants. The significance and evolution of these patterns are then discussed. Finally, we suggest several future directions of research, such as the exploration of temporal dynamics and the dissection of underlying mechanisms of virus-induced changes in whitefly behavior.

Screening of ulcerative colitis biomarkers and potential pathways based on weighted gene co-expression network, machine learning and ceRNA hypothesis.

BACKGROUND: Ulcerative colitis (UC) refers to an intractable intestinal inflammatory disease. Its increasing incidence rate imposes a huge burden on patients and society. The UC etiology has not been determined, so screening potential biomarkers is critical to preventing disease progression and selecting optimal therapeutic strategies more effectively. METHODS: The microarray datasets of intestinal mucosal biopsy of UC patients were selected from the GEO database, and integrated with R language to screen differentially expressed genes and draw proteins interaction network diagrams. GO, KEGG, DO and GSEA enrichment analyses were performed to explore their biological functions. Through machine learning and WGCNA analysis, targets that can be used as UC potential biomarkers are screened out. ROC curves were drawn to verify the reliability of the results and predicted the mechanism of marker genes from the aspects of immune cell infiltration, co-expression analysis, and competitive endogenous network (ceRNA). RESULTS: Two datasets GSE75214 and GSE87466 were integrated for screening, and a total of 107 differentially expressed genes were obtained. They were mainly related to biological functions such as humoral immune response and inflammatory response. Further screened out five marker genes, and found that they were associated with M0 macrophages, quiescent mast cells, M2 macrophages, and activated NK cells in terms of immune cell infiltration. The co-expression network found significant co-expression relationships between 54 miRNAs and 5 marker genes. According to the ceRNA hypothesis, NEAT1-miR-342-3p/miR-650-SLC6A14, NEAT1-miR-650-IRAK3, and XIST-miR-342-3p-IRAK3 axes were found as potential regulatory pathways in UC. CONCLUSION: This study screened out five biomarkers that can be used for the diagnosis and treatment of UC, namely SLC6A14, TIMP1, IRAK3, HMGCS2, and APOBEC3B. Confirmed that they play a role in the occurrence and development of UC at the level of immune infiltration, and proposed a potential RNA regulatory pathway that controls the progression of UC.

Enhanced Water Adsorption Performance of UiO-66 Modulated with p-Nitrobenzoic or p-Hydroxybenzoic Acid: Introduced Defects and Functional Groups.

Adding appropriate modulators can effectively improve the porosity and adsorption performance of UiO-66. Herein, UiO-66 samples were synthesized with p-nitrobenzoic acid (PNBA) and p-hydroxybenzoic acid (PHBA) as modulators. All samples exhibited good crystallinity and thermal stability. The polar functional groups (-NO2 and -OH) and defects were introduced into UiO-66, which significantly improved its water adsorption performance and applications in adsorption heat transformation. With the addition of six equiv PNBA, the saturated water uptake of UiO-66 increased from 0.40 to 0.58 g/g. Also, 4eqPNBA-UiO-66 exhibited the highest water uptake under low relative pressure, which was almost twice that of "low-defect" LD-UiO-66. The addition of PHBA had little effect on the saturated water absorption. However, its highest water uptake at P/P0 = 0.3 is 0.23 g/g, which is equivalent to that of 4eqPNBA-UiO-66. Ten consecutive adsorption/desorption cycles indicated that these samples had good cycle stability.

Comprehensive analysis of lysine lactylation in Frankliniella occidentalis.

Western flower thrips (Frankliniella occidentalis) are among the most important pests globally that transmit destructive plant viruses and infest multiple commercial crops. Lysine lactylation (Klac) is a recently discovered novel post-translational modification (PTM). We used liquid chromatography-mass spectrometry to identify the global lactylated proteome of F. occidentalis, and further enriched the identified lactylated proteins using Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO). In the present study, we identified 1,458 Klac sites in 469 proteins from F. occidentalis. Bioinformatics analysis showed that Klac was widely distributed in F. occidentalis proteins, and these Klac modified proteins participated in multiple biological processes. GO and KEGG enrichment analysis revealed that Klac proteins were significantly enriched in multiple cellular compartments and metabolic pathways, such as the ribosome and carbon metabolism pathways. Two Klac proteins were found to be involved in the regulation of the TSWV (Tomato spotted wilt virus) transmission in F. occidentalis. This study provides a systematic report and a rich dataset of lactylation in F. occidentalis proteome for potential studies on the Klac protein of this notorious pest.

Cross-Talk of Protein Expression and Lysine Acetylation in Response to TMV Infection in Nicotiana benthamiana.

Lysine acetylation (Kac), a reversible PTM, plays an essential role in various biological processes, including those involving metabolic pathways, pathogen resistance, and transcription, in both prokaryotes and eukaryotes. TMV, the major factor that causes the poor quality of Solanaceae crops worldwide, directly alters many metabolic processes in tobacco. However, the extent and function of Kac during TMV infection have not been determined. The validation test to detect Kac level and viral expression after TMV infection and Nicotinamide (NAM) treatment clarified that acetylation was involved in TMV infection. Furthermore, we comprehensively analyzed the changes in the proteome and acetylome of TMV-infected tobacco (Nicotiana benthamiana) seedlings via LC-MS/MS in conjunction with highly sensitive immune-affinity purification. In total, 2082 lysine-acetylated sites on 1319 proteins differentially expressed in response to TMV infection were identified. Extensive bioinformatic studies disclosed changes in acetylation of proteins engaged in cellular metabolism and biological processes. The vital influence of Kac in fatty acid degradation and alpha-linolenic acid metabolism was also revealed in TMV-infected seedlings. This study first revealed Kac information in N. benthamiana under TMV infection and expanded upon the existing landscape of acetylation in pathogen infection.

Proteome-wide analysis of lysine 2-hydroxyisobutyrylation in Frankliniella occidentalis.

BACKGROUND: Lysine 2-hydroxyisobutyrylation (Khib) is a novel and conserved post-translational modification (PTM). Frankliniella occidentalis are economically important agricultural pests globally and also notorious for vectoring destructive plant viruses. To better study the disease transmission mechanism of F. occidentalis, it is necessary to conduct in-depth analysis of it. So far, no Khib modification of insects has been reported. RESULTS: In this study, a proteome-wide analysis of Khib modifications in F. occidentalis was analyzed for the first time through the combination of high performance liquid chromatography fractionation technology and 2-hydroxyisobutyrylated peptide enrichment and other advanced technologies, 4093 Khib sites were identified on 1125 modified proteins. Bioinformatics and functional enrichment analyses showed that Khib-modified proteins were significantly enriched in many cell compartments and pathways, especially related to various cellular components and biological processes, and were more concentrated in ribosomes and proteasome subunits, involved in energy metabolism, protein synthesis and degradation, compared to the other nine species including Japonica rice, Homo sapiens, P. patens, Botrytis, Ustilaginoidea virens, Saccharomyces cerevisiae, T. gondii, C. albicans, and F. oxysporum. And Khib sites on virus-interacting insect proteins were discovered for the first time, such as cyclophilin and endoCP-GN. CONCLUSIONS: After three repeated experiments, we found a total of 4093 Khib sites on 1125 proteins. These modified proteins are mainly concentrated in ribosomes and proteasome subunits, and are widely involved in a variety of critical biological activities and metabolic processes of F. occidentalis. In addition, for the first time, Khib modification sites are found on the proteome of F. occidentalis, and these sites could be acted as for the virus interaction, including cyclophilin and endoCP-GN. The global map of 2-hydroxyisobutyrylation in thrips is an invaluable resource to better understand the biological processes of thrips and provide new means for disease control and mitigation of pest damage to crops.

Broad-spectrum chemicals block ROS detoxification to prevent plant fungal invasion.

Plant diseases cause a huge impact on food security and are of global concern. While application of agrochemicals is a common approach in the control of plant diseases currently, growing drug resistance and the impact of off-target effects of these compounds pose major challenges. The identification of pathogenicity-related virulence mechanisms and development of new chemicals that target these processes are urgently needed. One such virulence mechanism is the detoxification of reactive oxygen species (ROS) generated by host plants upon attack by pathogens. The machinery of ROS detoxification might therefore serve as a drug target for preventing plant diseases, but few anti-ROS-scavenging drugs have been developed. Here, we show that in the model system Botrytis cinerea secretion of the cytochrome c-peroxidase, BcCcp1 removes plant-produced H2O2 and promotes pathogen invasion. The peroxidase secretion is modulated by a Tom1-like protein, BcTol1, through physical interaction. We show that BcTol1 is regulated at different levels to enhance the secretion of BcCcp1 during the early infection stage. Inactivation of either BcTol1 or BcCcp1 leads to dramatically reduced virulence of B. cinerea. We identify two BcTol1-targeting small molecules that not only prevent B. cinerea invasion but also have effective activity against a wide range of plant fungal pathogens without detectable effect on the hosts. These findings reveal a conserved mechanism of ROS detoxification in fungi and provide a class of potential fungicides to control diverse plant diseases. The approach described here has wide implications for further drug discovery in related fields.

Field application of nanoliposomes delivered quercetin by inhibiting specific hsp70 gene expression against plant virus disease.

BACKGROUND: The annual economic loss caused by plant viruses exceeds 10 billion dollars due to the lack of ideal control measures. Quercetin is a flavonol compound that exerts a control effect on plant virus diseases, but its poor solubility and stability limit the control efficiency. Fortunately, the development of nanopesticides has led to new ideas. RESULTS: In this study, 117 nm quercetin nanoliposomes with excellent stability were prepared from biomaterials, and few surfactants and stabilizers were added to optimize the formula. Nbhsp70er-1 and Nbhsp70c-A were found to be the target genes of quercetin, through abiotic and biotic stress, and the nanoliposomes improved the inhibitory effect at the gene and protein levels by 33.6 and 42%, respectively. Finally, the results of field experiment showed that the control efficiency was 38% higher than that of the conventional quercetin formulation and higher than those of other antiviral agents. CONCLUSION: This research innovatively reports the combination of biological antiviral agents and nanotechnology to control plant virus diseases, and it significantly improved the control efficiency and reduced the use of traditional chemical pesticides.

Serratia marcescens-S3 inhibits Potato virus Y by activating ubiquitination of molecular chaperone proteins NbHsc70-2 in Nicotiana benthamiana.

The potato virus Y (PVY) is a plant virus that causes massive crop losses globally, especially in Solanaceae crops. A strain of the plant growth-promoting rhizobacterium (PGPR), Serratia marcescens-S3 was found to inhibit PVY replication in Nicotiana benthamiana. However, there have been no in-depth studies demonstrating the underlying mechanism. In the current study, we found that ubiquitination of NbHsc70-2 is an important way for Serratia marcescens-S3 to trigger induced systemic resistance (ISR). After the treatment with S. marcescens-S3, the protein level of NbHsc70-2 reduced significantly. Inhibiting of ubiquitination increased the accumulation of NbHsc70-2 in plants and reduced S. marcescens-S3-mediated resistance to PVY. Furthermore, transgenic engineered Nicotiana benthamiana NbHsc70-2KO and NbHsc70-2USM were constructed using CRISPR-Cas9-mediated NbHsc70-2 knock-out and ubiquitination respectively. S. marcescens-S3 significantly reduced the inhibition of NbHsc70-2 protein accumulation in NbHsc70-2KO and NbHsc70-2USM . The virulence of PVY was stronger in NbHsc70-2USM than the wild-type plants. These results showed that S. marcescens-S3 increases the ubiquitination of NbHsc70-2 to inhibit the recruitment of molecular chaperone NbHsc70-2 to reduce its replication and infection of PVY.