Author: Joana G. Vicente
Internal reviewer: Joël F. Pothier
Expert reviewer: Jessica L. Erickson

Class: XopL
Family: XopL
Prototype: XCV3220 (Xanthomonas euvesicatoria pv. euvesicatoria, ex Xanthomonas campestris pv. vesicatoria; strain 85-10)
GenBank ID: CAJ24951.1 (660 aa)
RefSeq ID: WP_218498620.1 (560 aa)
Examples of other sequences: XopL_Xcc306 21109412 (X. citri pv. citri); XopL_Xcc8004 66575899 (X. campestris pv. campestris)
3D structure: 4FC9, 4FCG (Singer et al., 2013). Full-length XopL_Xcv85-10 did not crystallize but fragments XopL[aa 144–450] and XopL[aa 474–660] yielded crystals (Singer et al., 2013). The crystal structure of the N-terminal region of XopL showed the presence of a leucine-rich repeat (LRR) domain, that might serve as a protein-protein interaction module for ubiquitination target recognition (Singer et al., 2013). The protein represents a new class of E3 ubiquitin ligases.

XopL was first identified in X. campestris pv. campestris CKGE_TMP_i ( CKGE_TMP_i Xcc CKGE_TMP_i ) strain 8004 as a candidate T3E due to the presence of a plant-inducible promoter (PIP) box upstream of the CDS, XC_4273 (Jiang CKGE_TMP_i et al. CKGE_TMP_i , 2009). The CDS CKGE_TMP_i XC_4273 CKGE_TMP_i , re-called XopXccLR (LR = leucine-rich repeat) or XopL_Xcc for the purposes of this article, in CKGE_TMP_i X CKGE_TMP_i . CKGE_TMP_i campestris CKGE_TMP_i pv. CKGE_TMP_i campestris CKGE_TMP_i 8004 was suggested to be a T3E as it harboured a N-terminal region possessing a translocation signal that was able to target proteins into plant cells (Jiang CKGE_TMP_i et al CKGE_TMP_i ., 2009). It was also shown to be required for CKGE_TMP_i X CKGE_TMP_i . CKGE_TMP_i campestris CKGE_TMP_i pv. CKGE_TMP_i campestris CKGE_TMP_i proliferation in hosts plant (Jiang CKGE_TMP_i et al CKGE_TMP_i ., 2009). It was only a few years later that the analysis of the genome sequence of CKGE_TMP_i Xcv CKGE_TMP_i strain 85-10 (synonymous with CKGE_TMP_i X. euvesicatoria CKGE_TMP_i 85-10; CKGE_TMP_i Xe CKGE_TMP_i 85-10) led to the identification of XCV3220 ( CKGE_TMP_i xopL_Xe CKGE_TMP_i ) as a new T3E candidate gene and to its more complete characterization (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013). This revealed XopLXe as an E3 ubiquitin ligase with a novel fold (XL-box), capable of interacting with the plant host ubiquitination cascade (Singer et al., 2023).

XopL_Xcc possesses features that are typical of T3Es: the promoter region of xopL_Xcc8004 gene contains a perfect plant inducible promoter (PIP) box followed by a 10 box similar sequence (TTCGC-N₁₅-TTCGC-N₃₁-ACGACA) and LRRs motif is characteristic of specific T3Es in pathogenic bacteria (Yan CKGE_TMP_i et al CKGE_TMP_i ., 2019). Using an AvrBs1 reporter fusion, XopL_Xcc was shown to be translated into plant cells in a CKGE_TMP_i hrpF CKGE_TMP_i - and CKGE_TMP_i hpaB CKGE_TMP_i -dependent manner (Jiang et al., 2009). XopL_Xe contains a PIP box (plant inducible promoter) in its promoter (TTCG-N₁₆-TTCG; genome position 3669238-261) and co-regulation with the T3S system was confirmed by RT-PCR (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013). Type III-dependent secretion and translocation was confirmed by CKGE_TMP_i in vitro CKGE_TMP_i secretion and CKGE_TMP_i in vivo CKGE_TMP_i translocation assays (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013).

The xopL _Xcc8004 gene contains a PIP box and was shown to be controlled by hrpG and hrpX (Jiang et al., 2009).

qRT-PCR revealed that transcript levels of 15 out of 18 tested non-TAL effector genes (as well as the regulatory genes hrpG and hrpX), including xopL, were significantly reduced in the Xanthomonas oryzae pv. oryzae ΔxrvC mutant compared with those in the wild-type strain PXO99^A (Liu et al., 2016).

The expression of xopL _Xcc8004 gene is positively regulated by HrpG/HrpX (Yan et al., 2019).

XopL_Xe from CKGE_TMP_i X. euvesicatoria CKGE_TMP_i 85-10 (previously referred to as CKGE_TMP_i X. campestris CKGE_TMP_i pv. CKGE_TMP_i euvesicatoria CKGE_TMP_i ; CKGE_TMP_i Xcv CKGE_TMP_i ) displays E3 ubiquitin ligase activity (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013).

XopL_Xe inhibits expression of the elf18- and flg22-induced defense gene pNHL10 in CKGE_TMP_i Arabidopsis CKGE_TMP_i mesophyll protoplasts independent of E3 ligase function (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013).

XopL_Xe suppresses ABA responsive reporter CKGE_TMP_i pRD29b:GUS CKGE_TMP_i and PTI reporter CKGE_TMP_i pFRK1:LUC CKGE_TMP_i in CKGE_TMP_i Arabidopsis CKGE_TMP_i protoplasts (Popov CKGE_TMP_i et al CKGE_TMP_i ., 2016).

XopL_Xcc interferes with innate immunity (Yan CKGE_TMP_i et al CKGE_TMP_i ., 2019; Huang et al., 2024a) and SA signaling in Arabidopsis (Huang et al., 2024).

XopL_Xe triggers cell death in CKGE_TMP_i Nicotiana benthamiana CKGE_TMP_i in an E3 ligase dependant manner (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013). XopL_Xoc from CKGE_TMP_i X. oryzae CKGE_TMP_i pv. CKGE_TMP_i oryzicola CKGE_TMP_i and XopL_Xoo from CKGE_TMP_i X. oryzae CKGE_TMP_i pv. CKGE_TMP_i oryzae CKGE_TMP_i PX099A also cause cell death in this model (Ma et al., 2020).

Distantly related XopL homolog XopL_Xcc from CKGE_TMP_i X. campestris CKGE_TMP_i pv. CKGE_TMP_i campestris CKGE_TMP_i 8004 failed to cause plant cell death. (Ortmann CKGE_TMP_i et al. CKGE_TMP_i , 2023).

XopL_Xcc is required for full virulence and growth of CKGE_TMP_i X. campestris CKGE_TMP_i pv. CKGE_TMP_i campestris CKGE_TMP_i 8004 in the host plant Chinese radish (Jiang CKGE_TMP_i et al. CKGE_TMP_i , 2009) but not in Arabidopsis (Huang et al, 2024a. However a mutant for 17 effectors (Δ CKGE_TMP_i 17E CKGE_TMP_i ) supplemented with CKGE_TMP_i xopL_Xcc CKGE_TMP_i did grow better than Δ CKGE_TMP_i 17E CKGE_TMP_i

XopL_Xcc constituative overexpression in Arabidopsis let to enhance Xcc 8004 virulence and suppressed callose deposition and oxidative burst (Huang et al., 2024)

XopL_Xe is required for full virulence of CKGE_TMP_i Xe CKGE_TMP_i 85-10 on tomato (Leong et al., 2022).

XopL_Xap supports CKGE_TMP_i X. axonopodis CKGE_TMP_i pv. CKGE_TMP_i punicae CKGE_TMP_i multiplication in pomegranate by suppressing plant immune responses including plant cell death (Soni CKGE_TMP_i et al CKGE_TMP_i ., 2017).

Transient expression of XopL_Xe, led to a nearly complete elimination of stromules and the relocation of plastids to the nucleus and further characterization revealed that the E3 ligase activity is essential for two plastid phenotypes (Erickson CKGE_TMP_i et al CKGE_TMP_i ., 2018).

CKGE_TMP_i Xcv CKGE_TMP_i strain 85-10 suppresses host autophagy by utilizing type-III effector XopL_Xe. Intriguingly, XopL_Xe is targeted for degradation by defense-related selective autophagy mediated by NBR1/Joka2, revealing a complex antagonistic interplay between XopL and the host autophagy machinery (Leong CKGE_TMP_i et al. CKGE_TMP_i , 2022).

Several localization patterns have been reported for XopL proteins in epidermal cells with some strain dependent differences. XopLs are most often tagged on the C-terminus, with the exception of the studies by Leong CKGE_TMP_i et al CKGE_TMP_i ., 2022 and Yan et al., 2019. Cytosolic localization has been reported for XopL_Xe (Erickson et al., 2018), XopL_Xcc (Yan et al., 2019; Ortmann et al. 2023) CKGE_TMP_i , CKGE_TMP_i XopL_Xoo (Ma et al., 2020; Ortmann et al. 2023; ), and XopL_Xac from CKGE_TMP_i X. citri CKGE_TMP_i pv. CKGE_TMP_i citri CKGE_TMP_i (Ortmann et al. 2023) in CKGE_TMP_i N. benthamiana CKGE_TMP_i . Nuclear localization was reported for XopL_Xe, XopL_Xcc (Yan CKGE_TMP_i et al. CKGE_TMP_i , 2019; Ortmann et al. 2023) and XopL_Xac in CKGE_TMP_i N. benthamiana CKGE_TMP_i (Ortmann et al. 2023), but not for XopL_Xoo in CKGE_TMP_i N. benthamiana CKGE_TMP_i or XopL_Xap from CKGE_TMP_i X. axonopodis CKGE_TMP_i pv CKGE_TMP_i punicae CKGE_TMP_i in CKGE_TMP_i A. thaliana CKGE_TMP_i protoplasts (Soni CKGE_TMP_i et al. CKGE_TMP_i , 2017; Ortmann et al. 2023). Plasma membrane localization has been reported for XopL_Xap transiently expressed in CKGE_TMP_i Nicotiana benthmiana CKGE_TMP_i (Soni CKGE_TMP_i et al CKGE_TMP_i ., 2017) and XopL CKGE_TMP_i _Xcc CKGE_TMP_i expressed in CKGE_TMP_i A. thaliana CKGE_TMP_i protoplasts (Huang CKGE_TMP_i et al. CKGE_TMP_i , 2024b; Yan CKGE_TMP_i et al. CKGE_TMP_i , 2019) and CKGE_TMP_i N. benthamiana CKGE_TMP_i leaves (Yan CKGE_TMP_i et al. CKGE_TMP_i , 2019). Microtubule localization has been reported for XopL_Xe, XopL_Xoo and XopL_Xac, whereas the distantly related XopL_Xcc failed to localize to microtubules (Ortmann CKGE_TMP_i et al. CKGE_TMP_i , 2023). Autophagosome localization has been repoted for XopL_Xe (co-localizes with autophagy markers RFP-ATG8e and SH3P2-RFP; Leong CKGE_TMP_i et al CKGE_TMP_i ., 2022).

XopL_Xe contains 9 leucine-rich repeats (LRRs). Mutation of amino acids in the central cavity of the XL-box disrupt E3 ligase activity and prevent XopL-induced plant cell death. The lack of cysteine residues in the XL-box suggest that thioester-linked ubiquitin-E3 ligase intermediates are not formed during XopL-mediated ubiquitination. Suppression of PAMP responses solely depends on the N-terminal LRR domain (Singer CKGE_TMP_i et al CKGE_TMP_i ., 2013).

XopL CKGE_TMP_i _Xe CKGE_TMP_i interacts with and degrades the autophagy component SH3P2 via its E3 ligase activity to promote infection (Leong CKGE_TMP_i et al. CKGE_TMP_i , 2022). XopL_Xoo interacts with and degrades ferredoxin from N. benthamiana, part of the electron transport chain (Ma et al., 2020). XopLXcc interaction with proton pump interactor 1 (PPI1) in Arabidopsis (Huang et al., 2024).

Yes (e.g., X. euvesicatoria, X. citri, X. axonopodis, X. oryzae, X. oryzicola, X. fragariae, X. perforans, X. gardneri, X. campestris pv. campestris, but not X. campestris pv. raphani, in some X. arboricola pathovars). See for example Table 2 in Jiang et al. (2009) and Figure S1 in Singer et al. (2013).

No.

Adlung N (2016). Charakterisierung der Avirulenzaktivität von XopQ und Identifizierung möglicher Interaktoren von XopL aus Xanthomonas campestris pv. vesicatoria. Doctoral Thesis. Martin-Luther-Universität Halle-Wittenberg, Germany. PDF: d-nb.info/1116951061/34

Erickson JL, Adlung N, Lampe C, Bonas U, Schattat MH (2018). The Xanthomonas effector XopL uncovers the role of microtubules in stromule extension and dynamics in Nicotiana benthamiana. Plant J. 93: 856-870. DOI:10.1111/tpj.13813

Huang J, Dong Y, Li N, He Y, Zhou H (2024). The type III effector XopL_Xcc in Xanthomonas campestris pv. campestris targets the proton pump interactor 1 and suppresses innate immunity in Arabidopsis. Int. J. Mol. Sci. 25: 9175. DOI: 10.3390/ijms25179175

Huang J, Zhou H, Zhou M, Li N, Jiang B, He Y (2024). Functional analysis of type III effectors in Xanthomonas campestris pv. campestris reveals distinct roles in modulating Arabidopsis innate immunity. Pathogens 13: 448. DOI: 10.3390/pathogens13060448

Jiang W, Jiang BL, Xu RQ, Huang JD, Wei HY, Jiang GF, Cen WJ, Liu J, Ge YY, Li GH, Su LL, Hang XH, Tang DJ, Lu GT, Feng JX, He YQ, Tang JL (2009). Identification of six type III effector genes with the PIP box in Xanthomonas campestris pv campestris and five of them contribute individually to full pathogenicity. Mol. Plant Microbe Interact. 22: 1401-1411. DOI: 10.1094/MPMI-22-11-1401

Leong JX, Raffeiner M, Spinti D, Langin G, Franz-Wachtel M, Guzman AR, Kim JG, Pandey P, Minina AE, Macek B, Hafrén A, Bozkurt TO, Mudgett MB, Börnke F, Hofius D, Üstün S (2022). A bacterial effector counteracts host autophagy by promoting degradation of an autophagy component. EMBO J. 41: e110352. DOI: 10.15252/embj.2021110352

Liu Y, Long J, Shen D, Song C (2016). Xanthomonas oryzae pv. oryzae requires H-NS-family protein XrvC to regulate virulence during rice infection. FEMS Microbiol. Lett. 363: fnw067. DOI: 10.1093/femsle/fnw067

Ma W, Xu X, Cai L, Cao Y, Haq F, Alfano JR, Zu B, Zou L, Chen G (2020) . A Xanthomonas oryzae type III effector XopL causes cell death through mediating ferredoxin degradation in Nicotiana benthamiana. Phytopathol Res. 2: 16. DOI: 10.1186/s42483-020-00055-w

Ortmann S, Marx J, Lampe C, Handrick V, Ehnert TM, Zinecker S, Reimers M, Bonas U, Erickson JL (2023). A conserved microtubule-binding region in Xanthomonas XopL is indispensable for induced plant cell death reactions. PLoS Pathog. 19: e1011263. DOI: 10.1371/journal.ppat.1011263

Popov G, Fraiture M, Brunner F, Sessa G (2016). Multiple Xanthomonas euvesicatoria type III effectors inhibit flg22-triggered immunity. Mol. Plant Microbe Interact. 29: 651-660. DOI: 10.1094/MPMI-07-16-0137-R

Singer AU, Schulze S, Skarina T, Xu X, Cui H, Eschen-Lippold L, Egler M, Srikumar T, Raught B, Lee J, Scheel D, Savchenko A, Bonas U (2013). A pathogen type III effector with a novel E3 ubiquitin ligase architecture. PLoS Pathog. 9: e1003121. DOI: 10.1371/journal.ppat.1003121

Soni M, Mondal KK. (2017). Xanthomonas axonopodis pv. punicae employs XopL effector to suppress pomegranate immunity. J. Integr. Plant Biol. 60: 341-357. DOI: 10.1111/jipb.12615

Yan X, Tao J, Luo HL, Tan LT, Rong W, Li HP, He CZ (2019). A type III effector XopL_Xcc8004 is vital for Xanthomonas campestris pathovar campestris to regulate plant immunity. Res. Microbiol. 170: 138-146. DOI: 10.1016/j.resmic.2018.12.001

This fact sheet is based upon work from COST Action CA16107 EuroXanth, supported by COST (European Cooperation in Science and Technology).