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bacteria:t3e:xopl

The Type III Effector XopL from //Xanthomonas//

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: XopLXcc306 21109412 (X. citri pv. citri); XopLXcc8004 66575899 (X. campestris pv. campestris)
3D structure: 4FC9, 4FCG (Singer et al., 2013). Full-length XopLXcv85-10 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.

Biological function

How discovered?

XopL was first identified in X. campestris pv. campestris (Xcc) strain 8004 as a candidate T3E due to the presence of a plant-inducible promoter (PIP) box upstream of the CDS, XC_4273 (Jiang et al., 2009). The CDS XC_4273, re-called XopXccLR (LR = leucine-rich repeat) or XopLXcc for the purposes of this article, in Xcc 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 et al., 2009). It was also shown to be required for Xcc proliferation in hosts plant (Jiang et al., 2009). It was only a few years later that the analysis of the genome sequence of X. campestris pv. vesicatoria (Xcv) strain 85-10 (synonymous with X. euvesicatoria 85-10; Xe 85-10) led to the identification of XCV3220 (xopLXe ) as a new T3E candidate gene and to its more complete characterization (Singer et al., 2013). XopLXe was shown to have E3 ubiquitin ligase activity, coferred by a ligase domain with a novel fold (XL-box), capable of interacting with the plant host ubiquitination cascade (Singer et al., 2023).

(Experimental) evidence for being a T3E

XopLXcc possesses features that are typical of T3Es: the promoter region of the xopLXcc gene contains a perfect plant inducible promoter (PIP) box followed by a 10 box similar sequence (TTCGC-N15-TTCGC-N31-ACGACA) and an LRR motif characteristic of T3Es in pathogenic bacteria (Yan et al., 2019). Using an AvrBs1 reporter fusion, XopLXcc was shown to be translocated into plant cells in a hrpF- and hpaB-dependent manner (Jiang et al., 2009). XopLXe also 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 et al., 2013). Type III-dependent secretion and translocation was confirmed by in vitro secretion and in vivo translocation assays (Singer et al., 2013).

Regulation

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 PXO99A (Liu et al., 2016).

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

Phenotypes

  • XopLXe from X. euvesicatoria 85-10 (previously referred to as X. campestris pv. euvesicatoria; Xcv) displays E3 ubiquitin ligase activity (Singer et al., 2013).
  • XopLXe inhibits expression of the elf18- and flg22-induced defense gene pNHL10 in Arabidopsis mesophyll protoplasts independent of E3 ligase function (Singer et al., 2013).
  • XopLXe suppresses ABA responsive reporter pRD29b:GUS and PTI reporter pFRK1:LUC in Arabidopsis protoplasts (Popov et al., 2016).
  • XopLXcc interferes with innate immunity (Yan et al., 2019; Huang et al., 2024a) and SA signaling in Arabidopsis (Huang et al., 2024).
  • XopLXe triggers cell death in Nicotiana benthamiana in an E3 ligase dependant manner (Singer et al., 2013). XopLXoc from X. oryzae pv. oryzicola and XopLXoo from X. oryzae pv. oryzae PX099A also cause cell death in this model (Ma et al., 2020).
  • Distantly related XopL homolog XopLXcc from Xcc 8004 failed to cause plant cell death. (Ortmann et al., 2023).
  • XopLXcc is required for full virulence and growth of Xcc 8004 in the host plant Chinese radish (Jiang et al., 2009) but not in Arabidopsis (Huang et al, 2024). However a mutant for 17 effectors (Δ17E) supplemented with xopLXcc did grow better than Δ17E.
  • XopLXcc constitutive overexpression in Arabidopsis led to enhanced Xcc 8004 virulence and suppressed callose deposition and oxidative burst (Huang et al., 2024).
  • XopLXe is required for full virulence of Xe 85-10 on tomato (Leong et al., 2022).
  • XopLXap supports X. axonopodis pv. punicae multiplication in pomegranate by suppressing plant immune responses including plant cell death (Soni et al., 2017).
  • Transient expression of XopLXe , 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 the two plastid phenotypes (Erickson et al., 2018).
  • Xe 85-10 suppresses host autophagy by utilizing type-III effector XopLXe . Intriguingly, XopLXe 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 et al., 2022).

Localization

Several localization patterns have been reported for XopL proteins in epidermal cells with some strain dependent differences. XopLs are most often tagged at the C-terminus, with the exception of the studies by Leong et al., 2022 and Yan et al., 2019.

  • Cytosolic localization has been reported for XopLXe (Erickson et al., 2018), XopLXcc (Yan et al., 2019; Ortmann et al. 2023), XopLXoo (Ma et al., 2020; Ortmann et al., 2023), and XopLXac from X. citri pv. citri (Ortmann et al. 2023) in N. benthamiana.
  • Nuclear localization was reported for XopLXe , XopLXcc (Yan et al., 2019; Ortmann et al., 2023) and XopLXac in N. benthamiana (Ortmann et al. 2023), but not for XopLXoo in N. benthamiana or XopLXap from X. axonopodis pv. punicae in Arabidopsis protoplasts (Soni et al., 2017; Ortmann et al., 2023).
  • Plasma membrane localization has been reported for XopLXap transiently expressed in N. benthmiana (Soni et al ., 2017) and XopLXcc expressed in Arabidopsis protoplasts (Huang et al. 2024b; Yan et al. , 2019) and N. benthamiana leaves (Yan et al., 2019).
  • Microtubule localization has been reported for XopLXe , XopLXoo and XopLXac , whereas the distantly related XopLXcc failed to localize to microtubules in N. benthamiana (Ortmann et al., 2023).
  • Autophagosome localization has been reported for XopLXe (co-localizes with autophagy markers RFP-ATG8e and SH3P2-RFP; Leong et al., 2022).

Enzymatic function

XopLXe harbours an unstructured N-terminus, followed by three alpha helices, an leucine-rich repeat (LRR) domain and an XL-box. 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 et al., 2013), while microtubule binding relies on a proline-rich region in the unstructured region and the alpha-helical region (Ortman et al., 2023).

Interaction partners

XopLXe interacts with and degrades the autophagy component SH3P2 via its E3 ligase activity to promote infection (Leong et al., 2022). XopLXoo 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., 2024b).

Conservation

In xanthomonads

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).

In other plant pathogens/symbionts

No.

References

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/1116951061/34FIXME

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 (2024a). The type III effector XopLXcc 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 (2024b). 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 XopLXcc8004 is vital for Xanthomonas campestris pathovar campestris to regulate plant immunity. Res. Microbiol. 170: 138-146. DOI: 10.1016/j.resmic.2018.12.001

Acknowledgements

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

bacteria/t3e/xopl.txt · Last modified: 2024/10/28 13:47 by jerickson