Transcription factor WRKY22 regulates canker susceptibility in sweet orange (Citrus sinensis Osbeck) by enhancing cell enlargement and CsLOB1 expression

Abstract

Pathological hypertrophy (cell enlargement) plays an important role in the development of citrus canker, but its regulators are largely unknown. Although WRKY22 is known to be involved in pathogen-triggered immunity and positively regulates resistance to bacterial pathogens in Arabidopsis, rice and pepper, the CRISPR/Cas9-mediated partial knockout of CsWRKY22 improves resistance to Xanthomonas citri subsp. citri (Xcc) in Wanjincheng orange (Citrus sinensis Osbeck). Here, we demonstrate that CsWRKY22 is a nucleus-localized transcriptional activator. CsWRKY22-overexpressing plants exhibited dwarf phenotypes that had wrinkled and thickened leaves and were more sensitive to Xcc, whereas CsWRKY22-silenced plants showed no visible phenotype changes and were more resistant to Xcc. Microscopic observations revealed that the overexpression of CsWRKY22 increased cell size in the spongy mesophyll. Transcriptome analysis showed that cell growth-related pathways, such as the auxin and brassinosteroid hormonal signaling and cell wall organization and biogenesis pathways, were significantly upregulated upon CsWRKY22 overexpression. Interestingly, CsWRKY22 activated the expression of CsLOB1, which is a key gene regulating susceptibility to citrus canker. We further confirmed that CsWRKY22 bound directly to the W-boxes just upstream of the transcription start site of CsLOB1 in vivo and in vitro. We conclude that CsWRKY22 enhances susceptibility to citrus canker by promoting host hypertrophy and CsLOB1 expression. Thus, our study provides new insights into the mechanism regulating pathological hypertrophy and the function of WRKY22 in citrus.

Overexpression of CsSAMT in Citrus sinensis Induces Defense Response and Increases Resistance to Xanthomonas citri subsp. citri

“…These two studies point to the involvement of SA levels and CsWRKY22 expression. In fact, these results corroborate our observation that plants overexpressing CsSAMT somehow repress CsWRKY22 and are more resistant to citrus canker disease ( Wang et al, 2019 ; Long et al, 2021 ). Curiously, the presence of the pathogen (T24) led to a decrease in expression of CsPAL , CsNPR1 , and CsPR1 defense-related genes in all the plants ( Supplementary Figure 8 ), but repression was transient in the transgenic plants.…”

“…Moreover, expression of NPR3 was higher only in VSAMT44 at T0, showing that the presence of free NPR1 monomers requires a high amount of NPR3, which is not observed in the presence of the pathogen (T24 and T48). Curiously, CsWRKY22 , a transcription factor associated with susceptibility to citrus canker and SA ( Kloth et al, 2016 ; Wang et al, 2019 ; Long et al, 2021 ), showed a significant lower expression in VSAMT44 at T0, when the pathogen was not present in the plant, suggesting that salicylic acid can modulate the expression of CsWRKY22 and consequently reflect the reduced symptoms observed in the primary infection ( Figure 6 ).…”

“…Thus, as NPR3 acts as a negative regulator of NPR1 ( Moreau et al, 2012 ), its overexpression in VSAMT44 could explain the normal phenotype observed in the transgenic lines, possibly due to the regulation of CsNPR1 levels by CsNPR3 . Repression of CsWRKY22 is directly associated with citrus canker resistance ( Wang et al, 2019 ; Long et al, 2021 ), and overexpression of CsWRKY22 in Arabidopsis mediated susceptibility to aphids, which is associated with the suppression of SA signaling ( Kloth et al, 2016 ). These two studies point to the involvement of SA levels and CsWRKY22 expression.…”

CmWRKY15-1 Promotes Resistance to Chrysanthemum White Rust by Regulating CmNPR1 Expression

“…CsWRKY22 , MdWRKY15 , and LiWRKY enhance the resistance of orange ( Citrus sinensis Osbeck), apple ( Malus × domestica ), and Easter lily ( Lilium longiflorum ) to Xanthomonas citri subsp. citri , Botryosphaeria dothidea , and Botrytis cinerea , respectively ( Zhao X. Y. et al, 2020 ; Kumari et al, 2021 ; Long et al, 2021 ). Heterologous expression of PtrWRKY73 in Arabidopsis ( Arabidopsis thaliana ) activates the expression of other downstream genes in the SA pathway and enhances resistance to the pathogen Pseudomonas syringae pathovar tomato strain DC3000 (Pst DC3000) ( Duan et al, 2015 ).…”

CsWRKY25 Improves Resistance of Citrus Fruit to Penicillium digitatum via Modulating Reactive Oxygen Species Production

“…In our previous study on citrus fruit, CsWRKY65 and CsWRKY70 were found to be important regulators of plant resistance against P. digitatum ( Deng et al, 2020 ; Wang et al, 2021 ). And CsWRKY22 in sweet orange could promote the expression of CsLOB1 and enhance the susceptibility to canker ( Long et al, 2021 ). Thus, the isolation and cloning of citrus disease resistance WRKY TFs genes and the exploration of molecular mechanism therein are of great significance.…”