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SUMMARY: Pustule formation is pivotal for the development of the Xanthomonas citri subsp. citri (Xcc)‐induced citrus canker disease (CCD). Although our previous study demonstrated that the exogenous application of abscisic acid (ABA) facilitated pustule formation induced by Xcc, the precise mechanism remains elusive. The 9‐cis‐epoxycarotenoid dioxygenase (NCED) is a crucial enzyme in ABA biosynthesis. This study explored the role of citrus CsNCED1‐1 in CCD resistance through overexpression and RNA interference of CsNCED1‐1 in Wanjincheng orange (Citrus sinensis). Our findings indicated that CsNCED1‐1 negatively modulated CCD resistance by fostering ABA accumulation, concomitant with an increase in jasmonic acid (JA) and a decrease in salicylic acid (SA). Plants overexpressing CsNCED1‐1 displayed shortened leaves with smaller and denser stomata along with irregular and increased palisade cells. CsLOB1 is a known susceptibility gene for CCD, and CsbZIP40 positively influences resistance to this disease. We further confirmed that CsLOB1 promoted and CsbZIP40 suppressed the transcription of CsNCED1‐1 by directly binding to the CsNCED1‐1 promoter. Notably, CsbZIP40 and CsLOB1 showed a competitive relationship in the regulation of CsNCED1‐1 expression, with CsbZIP40 exhibiting greater competitiveness. Overall, our findings highlight that CsNCED1‐1 promotes susceptibility to citrus canker by disrupting JA‐ and SA‐mediated defense mechanisms and triggering the proliferation and remodeling of palisade cells, thereby facilitating pathogen colonization and pustule formation. This study offers novel insights into the regulatory mechanisms underlying citrus canker resistance and the role of CsNCED1‐1 in citrus. [ABSTRACT FROM AUTHOR]

SUMMARY: Citrus production is severely threatened by the devastating Huanglongbing (HLB) disease globally. By studying and analyzing the defensive behaviors of an HLB‐tolerant citrus cultivar 'Shatangju', we discovered that citrus can sense Candidatus Liberibacter asiaticus (CLas) infection and induce immune responses against HLB, which can be further strengthened by both endogenously produced and exogenously applied methyl salicylate (MeSA). This immune circuit is turned on by an miR2977‐SAMT (encoding a citrus Salicylate [SA] O‐methyltransferase) cascade, by which CLas infection leads to more in planta MeSA production and aerial emission. We provided both transgenic and multi‐year trail evidences that MeSA is an effective community immune signal. Ambient MeSA accumulation and foliage application can effectively induce defense gene expression and significantly boost citrus performance. We also found that miRNAs are battle fields between citrus and CLas, and about 30% of the differential gene expression upon CLas infection are regulated by miRNAs. Furthermore, CLas hijacks host key processes by manipulating key citrus miRNAs, and citrus employs miRNAs that coordinately regulate defense‐related genes. Based on our results, we proposed that miRNAs and associated components are key targets for engineering or breeding resistant citrus varieties. We anticipate that MeSA‐based management, either induced expression or external application, would be a promising tool for HLB control. Significance Statement: The Great Wall not only served as a defensive barrier to ancient China, but also an intelligence transmission system. Foreign invasion alert at remote borders was signaled by rising smoke on the beacon towers and was relayed by the neighboring towers along the Great Wall. Recently researchers found a 'Great Wall' signaling system in citrus. Huanglongbing (HLB) is a devastating citrus disease causing billions of dollar losses annually. Research shows that citrus can produce and emit more methyl salicylate (MeSA) after being infected by CLas (the associated agent with HLB). MeSA is a violate that can spread through citrus growing groves and 'alarm' the uninfected citrus for the emerging threatens. This research also points out that microRNA (miRNA) is a key immune regulator in citrus. Image by: Hongwei Zhao. Due to our limited understanding of CLas pathogenicity and citrus immunity, citrus HLB causes enormous global economic losses yearly. In this investigation, we demonstrated that citrus can mount effective immune responses upon CLas infection, which is characterized by emitted community immune signal (MeSA) and citrus miRNA‐coordinated massive gene reprogramming. These two discoveries are of particular importance in the citrus industry. We could develop MeSA‐based HLB management strategies that would prime uninfected trees in the grove. In the future, we could also breed or engineer tolerant or even resistant citrus varieties and rootstocks relying on modifying critical citrus miRNAs. Our findings revealed not only citrus defensive tool kits against the HLB but also the potential of utilizing these weapons practically. [ABSTRACT FROM AUTHOR]

Summary: Citrus sinensis lateral organ boundary 1 (CsLOB1) was previously identified as a critical disease susceptibility gene for citrus bacterial canker, which is caused by Xanthomonas citri subsp. citri (Xcc). However, the molecular mechanisms of CsLOB1 in citrus response to Xcc are still elusive. Here, we constructed transgenic plants overexpressing and RNAi‐silencing of CsLOB1 using the canker‐disease susceptible 'wanjincheng' orange (C. sinensis Osbeck) as explants. CsLOB1‐overexpressing plants exhibited dwarf phenotypes with smaller and thicker leaf, increased branches and adventitious buds clustered on stems. These phenotypes were followed by a process of pustule‐ and canker‐like development that exhibited enhanced cell proliferation. Pectin depolymerization and expansin accumulation were enhanced by CsLOB1 overexpression, while cellulose and hemicellulose synthesis were increased by CsLOB1 silence. Whilst overexpression of CsLOB1 increased susceptibility, RNAi‐silencing of CsLOB1 enhanced resistance to canker disease without impairing pathogen entry. Transcriptome analysis revealed that CsLOB1 positively regulated cell wall degradation and modification processes, cytokinin metabolism, and cell division. Additionally, 565 CsLOB1‐targeted genes were identified in chromatin immunoprecipitation‐sequencing (ChIP‐seq) experiments. Motif discovery analysis revealed that the most highly overrepresented binding sites had a conserved 6‐bp 'GCGGCG' consensus DNA motif. RNA‐seq and ChIP‐seq data suggested that CsLOB1 directly activates the expression of four genes involved in cell wall remodeling, and three genes that participate in cytokinin and brassinosteroid hormone pathways. Our findings indicate that CsLOB1 promotes cell proliferation by mechanisms depending on cell wall remodeling and phytohormone signaling, which may be critical to citrus canker development and bacterial growth in citrus. [ABSTRACT FROM AUTHOR]