SDE19, a SEC-dependent effector from 'Candidatus Liberibacter asiaticus' suppresses plant immunity and targets Citrus sinensis Sec12 to interfere with vesicle trafficking.

Citrus huanglongbing (HLB), which is caused by the phloem-colonizing bacteria Candidatus Liberibacter asiaticus (CLas), poses a significant threat to citrus production worldwide. The pathogenicity mechanism of HLB remains poorly understood. SEC-dependent effectors (SDEs) have been suggested to play critical roles in the interaction between citrus and CLas. Here, we explored the function of CLIBASIA_05320 (SDE19), a core SDE from CLas, and its interaction with its host target. Our data revealed that SDE19 is expressed at higher level during infection of citrus than that during infection of the Asian citrus psyllid. Subcellular localization assays showed that SDE19 is localized in the nucleus and cytoplasm and is capable of moving from cell to cell in Nicotiana benthamiana. To investigate whether SDE19 facilitates pathogen infection, we generated transgenic Arabidopsis thaliana and citrus plants overexpressing SDE19. Transgenic A. thaliana and citrus plants were more susceptible to Pseudomonas syringae pv. tomato (Pst) and Xanthomonas citri subsp. citri (Xcc), respectively. In addition, RNA-seq analysis demonstrated that overexpression of SDE19 resulted in a reprogramming of expression of genes related to biotic stimulus responses. SDE19 interacts with Citrus sinensis Sec12, a guanine nucleotide exchange factor responsible for the assembly of plant COPII (coat protein II)-coated vesicles, which mediate vesicle trafficking from the ER to the Golgi. SDE19 colocalizes with Sec12 in the ER by binding to its N-terminal catalytic region, affecting the stability of Sec12 through the 26S proteasome. This interaction hinders the secretion of apoplastic defense-related proteins such as PR1, P69B, GmGIP1, and RCR3. Furthermore, the secretion of PR1 and callose deposition is decreased in SDE19-transgenic A. thaliana. Taken together, SDE19 is a novel virulent SDE secreted by CLas that interacts with Sec12 to disrupt vesicle trafficking, inhibit defense-related proteins secretion, and promote bacterial infection. This study sheds light on how CLas manipulates the host vesicle trafficking pathway to suppress the secretion of defense-related proteins and interfere with plant immunity.

OrangeExpDB: an integrative gene expression database for Citrus spp.

BACKGROUND: Citrus is a major fruit crop, and RNA-sequencing (RNA-seq) data can be utilized to investigate its gene functions, heredity, evolution, development, and the detection of genes linked to essential traits or resistance to pathogens. However, it is challenging to use the public RNA-seq datasets for researchers without bioinformatics training, and expertise. RESULTS: OrangeExpDB is a web-based database that integrates transcriptome data of various Citrus spp., including C. limon (L.) Burm., C. maxima (Burm.) Merr., C. reticulata Blanco, C. sinensis (L.) Osbeck, and Poncirus trifoliata (L.) Raf., downloaded from the NCBI SRA database. It features a blast tool for browsing and searching, enabling quick download of expression matrices for different transcriptome samples. Expression of genes of interest can be easily generated by searching gene IDs or sequence similarity. Expression data in text format can be downloaded and presented as a heatmap, with additional sample information provided at the bottom of the webpage. CONCLUSIONS: Researchers can utilize OrangeExpDB to facilitate functional genomic analysis and identify key candidate genes, leveraging publicly available citrus RNA-seq datasets. OrangeExpDB can be accessed at http://www.orangeexpdb.com/ .

Phytophthora infestans RXLR effector PITG20303 targets a potato MKK1 protein to suppress plant immunity.

Pathogens secret a plethora of effectors into the host cell to modulate plant immunity. Analysing the role of effectors in altering the function of their host target proteins will reveal critical components of the plant immune system. Here we show that Phytophthora infestans RXLR effector PITG20303, a virulent variant of AVRblb2 (PITG20300) that escapes recognition by the resistance protein Rpi-blb2, suppresses PAMP-triggered immunity (PTI) and promotes pathogen colonization by targeting and stabilizing a potato MAPK cascade protein, StMKK1. Both PITG20300 and PITG20303 target StMKK1, as confirmed by multiple in vivo and in vitro assays, and StMKK1 was shown to be a negative regulator of plant immunity, as determined by overexpression and gene silencing. StMKK1 is a negative regulator of plant PTI, and the kinase activities of StMKK1 are required for its suppression of PTI and effector interaction. PITG20303 depends partially on MKK1, PITG20300 does not depend on MKK1 for suppression of PTI-induced reactive oxygen species burst, while the full virulence activities of nuclear targeted PITG20303 and PITG20300 are dependent on MKK1. Our results show that PITG20303 and PITG20300 target and stabilize the plant MAPK cascade signalling protein StMKK1 to negatively regulate plant PTI response.

The novel peptide NbPPI1 identified from Nicotiana benthamiana triggers immune responses and enhances resistance against Phytophthora pathogens.

In plants, recognition of small secreted peptides, such as damage/danger-associated molecular patterns (DAMPs), regulates diverse processes, including stress and immune responses. Here, we identified an SGPS (Ser-Gly-Pro-Ser) motif-containing peptide, Nicotiana tabacum NtPROPPI, and its two homologs in Nicotiana benthamiana, NbPROPPI1 and NbPROPPI2. Phytophthora parasitica infection and salicylic acid (SA) treatment induced NbPROPPI1/2 expression. Moreover, SignalP predicted that the 89-amino acid NtPROPPI includes a 24-amino acid N-terminal signal peptide and NbPROPPI1/2-GFP fusion proteins were mainly localized to the periplasm. Transient expression of NbPROPPI1/2 inhibited P. parasitica colonization, and NbPROPPI1/2 knockdown rendered plants more susceptible to P. parasitica. An eight-amino-acid segment in the NbPROPPI1 C-terminus was essential for its immune function and a synthetic 20-residue peptide, NbPPI1, derived from the C-terminus of NbPROPPI1 provoked significant immune responses in N. benthamiana. These responses led to enhanced accumulation of reactive oxygen species, activation of mitogen-activated protein kinases, and up-regulation of the defense genes Flg22-induced receptor-like kinase (FRK) and WRKY DNA-binding protein 33 (WRKY33). The NbPPI1-induced defense responses require Brassinosteroid insensitive 1-associated receptor kinase 1 (BAK1). These results suggest that NbPPI1 functions as a DAMP in N. benthamiana; this novel DAMP provides a potentially useful target for improving plant resistance to Pytophthora pathogens.

Negative regulators of plant immunity derived from cinnamyl alcohol dehydrogenases are targeted by multiple Phytophthora Avr3a-like effectors.

Oomycete pathogens secrete numerous effectors to manipulate host immunity. While some effectors share a conserved structural fold, it remains unclear if any have conserved host targets. Avr3a-like family effectors, which are related to Phytophthora infestans effector PiAvr3a and are widely distributed across diverse clades of Phytophthora species, were used to study this question. By using yeast-two-hybrid, bimolecular fluorescence complementation and co-immunoprecipitation assays, we identified members of the plant cinnamyl alcohol dehydrogenase 7 (CAD7) subfamily as targets of multiple Avr3a-like effectors from Phytophthora pathogens. The CAD7 subfamily has expanded in plant genomes but lost the lignin biosynthetic activity of canonical CAD subfamilies. In turn, we identified CAD7s as negative regulators of plant immunity that are induced by Phytophthora infection. Moreover, AtCAD7 was stabilized by Avr3a-like effectors and involved in suppression of pathogen-associated molecular pattern-triggered immunity, including callose deposition, reactive oxygen species burst and WRKY33 expression. Our results reveal CAD7 subfamily proteins as negative regulators of plant immunity that are exploited by multiple Avr3a-like effectors to promote infection in different host plants.

Analysis of huanglongbing-associated RNA-seq data reveals disturbances in biological processes within Citrus spp. triggered by Candidatus Liberibacter asiaticus infection.

Introduction: Huanglongbing (HLB), a disease that's ubiquitous worldwide, wreaks havoc on the citrus industry. The primary culprit of HLB is the gram-negative bacterium Candidatus Liberibacter asiaticus (CLas) that infects the phloem, but its damaging mechanism is yet to be fully understood. Methods and results: In this study, a multitude of tools including weighted correlation network analysis (WGCNA), protein-protein interaction (PPI) network analysis and gene expression profiling are employed to unravel the intricacies of its pathogenesis. The investigation pinpoints various central genes, such as the ethylene-responsive transcription factor 9 (ERF9) and thioredoxin reductase 1 (TrxR1), that are associated with CLas invasion and resultant disturbances in numerous biological operations. Additionally, the study uncovers a range of responses through the detection of differential expressed genes (DEGs) across different experiments. The discovery of core DEGs leads to the identification of pivotal genes such as the sieve element occlusion (SEO) and the wall-associated receptor kinase-like 15 (WAKL15). PPI network analysis highlights potential vital proteins, while GO and KEGG pathway enrichment analysis illustrate a significant impact on multiple defensive and metabolic pathways. Gene set enrichment analysis (GSEA) indicates significant alterations in biological processes such as leaf senescence and response to biotic stimuli. Discussion: This all-encompassing approach extends valuable understanding into the pathogenesis of CLas, potentially aiding future research and therapeutic strategies for HLB.

Transcriptome analysis of Citrus sinensis reveals potential responsive events triggered by Candidatus Liberibacter asiaticus.

Citrus Huanglongbing (HLB), caused by Candidatus Liberibacter asiaticus (CLas), is a devastating immune-mediated disorder that has a detrimental effect on the citrus industry, with the distinguishing feature being an eruption of reactive oxygen species (ROS). This study explored the alterations in antioxidant enzyme activity, transcriptome, and RNA editing events of organelles in C. sinensis during CLas infection. Results indicated that there were fluctuations in the performance of antioxidant enzymes, such as ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), peroxidase (POD), and superoxide dismutase (SOD), in plants affected by HLB. Transcriptome analysis revealed 3604 genes with altered expression patterns between CLas-infected and healthy samples, including those associated with photosynthesis, biotic interactions, and phytohormones. Samples infected with CLas showed a decrease in the expression of most genes associated with photosynthesis and gibberellin metabolism. It was discovered that RNA editing frequency and the expression level of various genes in the chloroplast and mitochondrion genomes were affected by CLas infection. Our findings provide insights into the inhibition of photosynthesis, gibberellin metabolism, and antioxidant enzymes during CLas infection in C. sinensis.

Genome-wide characterization of heavy metal-associated isoprenylated plant protein gene family from Citrus sinensis in response to huanglongbing.

Introduction: Heavy metal-associated isoprenylated plant proteins (HIPPs) play vital roles in maintaining heavy metal balance and responding to both biotic and abiotic stresses in vascular plants. However, the role of HIPPs in the response to Huanglongbing (HLB), a harmful disease of citrus caused by the phloem-colonizing bacterium Candidatus Liberibacter asiaticus (CLas), has not been examined. Methods and results: In this study, a total of 26 HIPP genes were identified in Citrus sinensis, and they were grouped into 5 clades. The CsHIPP genes are distributed on 8 chromosomes and exhibited considerable synteny with HIPPs found in Arabidopsis thaliana. Additionally, we analyzed the gene structure, conserved motifs and domains of the CsHIPPs. Various cis-acting elements related to plant hormones and stress responses were identified in the promoters of CsHIPPs. Public transcriptome data and RT-qPCR analysis showed that the expression level of CsHIPP03 was significantly reduced in samples infected by CLas and Xanthomonas citri ssp. citri (Xcc). Furthermore, silencing the homologous gene of CsHIPP03 in Nicotiana benthamiana increased the disease resistance of plants to bacteria. Discussion: Our results provide a basis for functional studies of HIPP gene family in C. sinensis, highlighting their functions in bacterial resistance, and improve our understanding to the susceptibility mechanism of HLB.

Synthesis and Properties of Room-Temperature Phosphorescent Liquid Crystal Copolymers with Linearly Polarized Luminescence Characteristic.

Room-temperature phosphorescent (RTP) liquid crystal materials have garnered considerable attention because of their significant applications in organic light emitting diodes, polarized light emitting materials, and so forth. How to efficiently synthesize pure organic RTP liquid crystals and regulate their performance is of great significance. In this article, we propose a simple and feasible method to synthesize RTP liquid crystals and manipulate their properties through copolymerization. We constructed RTP liquid crystal copolymers by copolymerizing a phosphorescent monomer bearing biphenyl mesogen with a phosphorescent monomer bearing a dibenzofuran chromophore. All the synthesized copolymers show a liquid crystal property because of the introduction of biphenyl mesogen. Meanwhile, by changing the composition of copolymers, it is possible to regulate their RTP performance, including luminescence color and lifetime. As the content of the PMDFM0C component in copolymers increases, the phosphorescence lifetime gradually increases. For poly(MDFM0C(0.46)-co-MBi18C(0.54)), the phosphorescence lifetime can reach 463.0 ms. Moreover, the phosphorescence color of the PMDFM0C component in copolymers changes with the copolymer composition, which can induce variable room-temperature phosphorescence. In addition, when oriented, liquid crystal copolymer films can emit linearly polarized fluorescence and linearly polarized phosphorescence. The linearly polarized phosphorescence dichroic ratio and polarization ratio values of the oriented poly(MDFM0C(0.46)-co-MBi18C(0.54)) film are 3.33 and 0.50, respectively.