The CsAP2-09-CsWRKY25-CsRBOH2 cascade confers resistance against citrus bacterial canker by regulating ROS homeostasis.

Citrus bacterial canker (CBC) is a serious bacterial disease caused by Xanthomonas citri subsp. citri (Xcc) that adversely impacts the global citrus industry. In a previous study, we demonstrated that overexpression of an Xcc-inducible apetala 2/ethylene response factor encoded by Citrus sinensis, CsAP2-09, enhances CBC resistance. The mechanism responsible for this effect, however, is not known. In the present study, we showed that CsAP2-09 targeted the promoter of the Xcc-inducible WRKY transcription factor coding gene CsWRKY25 directly, activating its transcription. CsWRKY25 was found to localize to the nucleus and to activate transcriptional activity. Plants overexpressing CsWRKY25 were more resistant to CBC and showed higher expression of the respiratory burst oxidase homolog (RBOH) CsRBOH2, in addition to exhibiting increased RBOH activity. Transient overexpression assays in citrus confirmed that CsWRKY25 and CsRBOH2 participated in the generation of reactive oxygen species (ROS) bursts, which were able to restore the ROS degradation caused by CsAP2-09 knockdown. Moreover, CsWRKY25 was found to bind directly to W-box elements within the CsRBOH2 promoter. Notably, CsRBOH2 knockdown had been reported previously to reduce the CBC resistance, while demonstrated in this study, CsRBOH2 transient overexpression can enhance the CBC resistance. Overall, our results outline a pathway through which CsAP2-09-CsWRKY25 transcriptionally reprograms CsRBOH2-mediated ROS homeostasis in a manner conducive to CBC resistance. These data offer new insight into the mechanisms and regulatory pathways through which CsAP2-09 regulates CBC resistance, highlighting its potential utility as a target for the breeding of CBC-resistant citrus varieties.

The wall-associated receptor-like kinase CsWAKL01, positively regulated by the transcription factor CsWRKY53, confers resistance to citrus bacterial canker via regulation of phytohormone signaling.

Citrus bacterial canker (CBC) is a disease that poses a major threat to global citrus production and is caused by infection with Xanthomonas citri subsp. citri (Xcc). Wall-associated receptor-like kinase (WAKL) proteins play an important role in shaping plant resistance to various bacterial and fungal pathogens. In a previous report, CsWAKL01 was identified as a candidate Xcc-inducible gene found to be up-regulated in CBC-resistant citrus plants. However, the functional role of CsWAKL01 and the mechanisms whereby it may influence resistance to CBC have yet to be clarified. Here, CsWAKL01 was found to localize to the plasma membrane, and the overexpression of the corresponding gene in transgenic sweet oranges resulted in pronounced enhancement of CBC resistance, whereas its knockdown had the opposite effect. Mechanistically, the effect of CsWAKL01 was linked to its ability to reprogram jasmonic acid, salicylic acid, and abscisic acid signaling activity. CsWRKY53 was further identified as a transcription factor capable of directly binding to the CsWAKL01 promoter and inducing its transcriptional up-regulation. CsWRKY53 silencing conferred greater CBC susceptibility to infected plants. Overall, these data support a model wherein CsWRKY53 functions as a positive regulator of CsWAKL01 to enhance resistance to CBC via the reprogramming of phytohormone signaling. Together these results offer new insights into the mechanisms whereby WAKLs shape phytopathogen resistance while underscoring the potential value of targeting the CsWRKY53-CsWAKL01 axis when seeking to breed CBC-resistant citrus plant varieties.

Genome-Wide Identification and Characterization of the Sweet Orange (Citrus sinensis) GATA Family Reveals a Role for CsGATA12 as a Regulator of Citrus Bacterial Canker Resistance.

Citrus bacterial canker (CBC) is a severe bacterial infection caused by Xanthomonas citri subsp. citri (Xcc), which continues to adversely impact citrus production worldwide. Members of the GATA family are important regulators of plant development and regulate plant responses to particular stressors. This report aimed to systematically elucidate the Citrus sinensis genome to identify and annotate genes that encode GATAs and evaluate the functional importance of these CsGATAs as regulators of CBC resistance. In total, 24 CsGATAs were identified and classified into four subfamilies. Furthermore, the phylogenetic relationships, chromosomal locations, collinear relationships, gene structures, and conserved domains for each of these GATA family members were also evaluated. It was observed that Xcc infection induced some CsGATAs, among which CsGATA12 was chosen for further functional validation. CsGATA12 was found to be localized in the nucleus and was differentially upregulated in the CBC-resistant and CBC-sensitive Kumquat and Wanjincheng citrus varieties. When transiently overexpressed, CsGATA12 significantly reduced CBC resistance with a corresponding increase in abscisic acid, jasmonic acid, and antioxidant enzyme levels. These alterations were consistent with lower levels of salicylic acid, ethylene, and reactive oxygen species. Moreover, the bacteria-induced CsGATA12 gene silencing yielded the opposite phenotypic outcomes. This investigation highlights the important role of CsGATA12 in regulating CBC resistance, underscoring its potential utility as a target for breeding citrus varieties with superior phytopathogen resistance.

Genome-wide identification and characterization of the sweet orange (Citrus sinensis) basic helix-loop-helix (bHLH) family reveals a role for CsbHLH085 as a regulator of citrus bacterial canker resistance.

Citrus bacterial canker (CBC) is a harmful bacterial disease caused by Xanthomonas citri subsp. citri (Xcc), negatively impacting citrus production worldwide. The basic helix-loop-helix (bHLH) transcription factor family plays crucial roles in plant development and stress responses. This study aimed to identify and annotate bHLH proteins encoded in the Citrus sinensis genome and explore their involvement and functional importance in regulating CBC resistance. A total of 135 putative CsbHLHs TFs were identified and categorized into 16 subfamilies. Their chromosomal locations, collinearity, and phylogenetic relationships were comprehensively analyzed. Upon Xcc strain YN1 infection, certain CsbHLHs were differentially regulated in CBC-resistant and CBC-sensitive citrus varieties. Among these, CsbHLH085 was selected for further functional characterization. CsbHLH085 was upregulated in the CBC-resistant citrus variety, was localized in the nucleus, and had a transcriptional activation activity. CsbHLH085 overexpression in Citrus significantly enhanced CBC resistance, accompanied by increased levels of salicylic acid (SA), jasmonic acid (JA), reactive oxygen species (ROS), and decreased levels of abscisic acid (ABA) and antioxidant enzymes. Conversely, CsbHLH085 virus-induced gene silencing resulted in opposite phenotypic and biochemical responses. CsbHLH085 silencing also affected the expression of phytohormone biosynthesis and signaling genes involved in SA, JA, and ABA signaling. These findings highlight the crucial role of CsbHLH085 in regulating CBC resistance, suggesting its potential as a target for biotechnological-assisted breeding citrus varieties with improved resistance against phytopathogens.

Systematic analysis and functional verification of citrus ascorbate peroxidases reveal that CsAPX01 and CsAPX02 negatively regulate citrus bacterial canker through the hydrogen peroxide regulation.

Ascorbate peroxidases (APXs) are antioxidant enzymes that play vital roles in redox homeostasis in plants. Citrus is susceptible to infection by Xanthomonas citri subsp. citri (Xcc), resulting in citrus bacterial canker (CBC). The present study used bioinformatic and expression analyses to investigate the APX family in Citrus sinensis. Bioinformatic research revealed the chromosomal locations, phylogeny, gene structure, promoter elements, functional domains, conserved motifs, and most likely physicochemical properties of the sequences. Six APXs clustered in three groups were identified, with each protein containing a single peroxidase domain. The promoter regions contained a variety of transcription factor-binding and hormone-response components. Xcc infection induced different CsAPX01 and CsAPX02 expressions in the CBC-susceptible Wanjincheng and CBC-resistant Kumquat varieties. Subcellular localization and transient expression showed that CsAPX01 and CsAPX02 were expressed in the cytoplasm and nucleus and had hydrogen peroxide (H2O2)-scavenging activity. Virus-induced gene silencing (VIGS) of CsAPX01 and CsAPX02 resulted in strong resistance to CBC and H2O2 bursts without effects on the plant phenotype. The current study focused on investigating and characterizing the citrus APX family. It was found that CsAPX01 and CsAPX02 exacerbated CBC by altering the balance of H2O2. These findings emphasize the importance of APXs in enhancing plant resistance to pathogens.

Rational construction of Co-promoted 1T-MoS2 nanoflowers towards high-efficiency 4-nitrophenol reduction.

Developing efficient and cost-effective non-noble metal catalysts for the reduction of 4-nitrophenol (4-NP) to 4-aminophenol (4-AP) is of great importance. Herein, Co-promoted 1T-MoS2 nanoflowers were synthesized via a one-step hydrothermal method. The influence of Co content on the structure and catalytic performance of 1T-MoS2 was studied in detail. It was found that Co doping not only enhanced the electronic conductivity but also increased the hydrogen adsorption ability of 1T-MoS2. Meanwhile, the highest activity was achieved due to the synergy effect of Co-Mo-S and CoS2 active phase. In the catalytic reduction of 4-NP, the reaction rate constant of Co/1T-MoS2-0.3 was as high as 0.908 min-1 and the catalyst exhibited excellent stability after recycling five times. The present work provides new insights for the rational design of highly efficient metal-doped MoS2 catalysts towards 4-NP reduction in wastewater.

MDTL-ACP: Anticancer Peptides Prediction Based on Multi-Domain Transfer Learning.

Anticancer peptides (ACPs) have emerged as one of the most promising therapeutic agents for cancer treatment. They are bioactive peptides featuring broad-spectrum activity and low drug-resistance. The discovery of ACPs via traditional biochemical methods is laborious and costly. Accordingly, various computational methods have been developed to facilitate the discovery of ACPs. However, the data resources and knowledge of ACPs are still very scarce, and only a few of them are clinically verified, which limits the competence of computational methods. To address this issue, in this paper, we propose an ACP prediction model based on multi-domain transfer learning, namely MDTL-ACP, to discriminate novel ACPs from plentiful inactive peptides. In particular, we collect abundant antimicrobial peptides (AMPs) from four well-studied peptide domains and extract their inherent features as the input of MDTL-ACP. The features learned from multiple source domains of AMPs are then transferred into the target prediction task of ACPs via artificial neural network-based shared-extractor and task-specific classifiers in MDTL-ACP. The knowledge captured in the transferred features enhances the prediction of ACPs in the target domain. Experimental results demonstrate that MDTL-ACP can outperform the traditional and state-of-the-art ACP prediction methods. The source code of MDTL-ACP and the data used in this study are available at https://github.com/JunhangCao/MTL-ACP.

CsAP2-09 confers resistance against citrus bacterial canker by regulating CsGH3.1L-mediated phytohormone biosynthesis.

Citrus bacterial canker (CBC) is a serious bacterial disease affecting citrus plantations and the citrus industry all over the world. We have previously shown that an apetala 2/ethylene response factor in Citrus sinensis, CsAP2-09, positively regulated resistance to CBC, although the regulatory mechanisms remained undetermined. Here, we demonstrated that CsAP2-09 positively and sustainably controlled resistance to CBC in three-year transgenic plants. CsAP2-09 was found to be a transcriptional activator, and qRT-PCR and dual luciferase assays showed that it controlled the expression CsGH3.1L. CsAP2-09 bound directly to the promotor of CsGH3.1L, shown by yeast one-hybrid assay, with the binding site confirmed by electrophoretic mobility shift assay. Biochemical assays showed that CsAP2-09 negatively regulated the biosynthesis of indole acetic acid (IAA) and positively regulated that of salicylic acid (SA) and ethylene, verified with transient overexpression of CsGH3.1L. The combination of these results with those of previous reports indicated that SA, ethylene, and IAA can directly regulate CBC resistance. Overall, we revealed a pathway whereby CsAP2-09 conferred CBC resistance by direct binding to the CsGH3.1L promoter, activating its expression and modulating IAA, SA, and ethylene biosynthesis. Our study indicates the potential value of manipulating CsAP2-09 and CsGH3.1L in the breeding of CBC-resistant citrus.

CsBZIP40 confers resistance against citrus bacterial canker by repressing CsWRKY43-CsPrx53/CsSOD13 cascade mediated ROS scavenging.

As the bacterial etiologic agent causing citrus bacterial canker (CBC), Xanthomonas citri subsp. citri (Xcc) seriously impacts citrus plantation and fruit production globally. In an earlier study, we demonstrated that CsBZIP40 can positively impact CBC resistance in the sweet orange (Citrus sinensis). However, the mechanistic basis for the protective benefits conferred by CsBZIP40 is yet to be delineated. Here, we show that CsBZIP40 positively regulates CBC resistance and reactive oxygen species (ROS) homeostasis in transgenic sweet orange overexpressing CsBZIP40. CsBZIP40 directly binds to the TGA-box of the CsWRKY43 promoter to repress its transcriptional activity. CsWRKY43 overexpression induces CBC susceptibility in transgenic sweet oranges. In contrast, its inhibition produces strong resistance to CBC. CsWRKY43 directly binds to the W-boxes of the CsPrx53 and CsSOD13 promoters to positively regulate the activities of these antioxidant enzymes, resulting in the negative regulation of ROS homeostasis and CBC resistance in sweet orange plants. CsPrx53/CsSOD13 knockdown enhances ROS accumulation and CBC resistance. Overall, our results outline a regulatory pathway through which CsBZIP40 transcriptionally represses CsWRKY43-CsPrx53/CsSOD13 cascade-mediated ROS scavenging in a manner conducive to CBC resistance. These mechanisms underscore the potential importance of CsBZIP40, CsWRKY43, CsPrx53, and CsSOD13, providing promising strategies for the prevention of CBC.