Author: Saul Burdman
Internal reviewer: Joël F. Pothier
Expert reviewer: Doron Teper

Class: XopAP
Family: XopAP
Prototype: XPE_1567 (Xanthomonas euvesicatoria pv. perforans; strain 91- 118) and XCV3138 (Xanthomonas euvesicatoria pv. euvesicatoria, ex Xanthomonas campestris pv. vesicatoria; strain 85-10)
GenBank ID (XPE_1567): AQS78154.1 (358 aa)
GenBank ID (XCV3138): CAJ24869.1 (464 aa)
RefSeq ID: WP_236504155.1 (426 aa)
3D structure: Unknown

XopAO was predicted to be a type 3 effector based on homology to Rip38, a predicted type 3 effector from Ralstonia solanacearum (Potnis et al., 2011). XopAP (XCV3138 in X. euvesicatoria strain 85-10; GenBank AM039952.1) was re-discovered using a machine-learning approach (Teper et al., 2016).

XopAP fused to the AvrBs2 reporter was shown to translocate into plant cells in an hrpF-dependent manner (Teper et al., 2016).

The xopAP gene was shown to be induced in X. citri subsp. citri strain 306 in nutrient broth (Jalan et al., 2013). In X. euvesicatoria strain 85-10, the xopAP gene does not contain a PIP-box motif in its promoter region (Teper et al., 2016). xopAP in X. citri pv. citri is positively regulated by the stringent response regulators RelA and SpoT (Zhang et al. 2019).

A Xanthomonas euvesicatoria strain 85-10 mutant defective in xopAP was compromised in induction of disease symptoms in leaves of susceptible pepper plants, relative to wild-type 85-10. This phenotype was associated with reduced ion leakage and higher chlorophyll content as compared with leaves inoculated with wild-type 85-10. No differences were observed between the xopAP mutant and wild-type 85-10 in their ability to colonize the leaves of susceptible pepper plants (Teper et al., 2016). Agrobacterium-mediated expression of XopAP in Nicotiana benthamiana caused a bleaching phenotype that was detected three days after agroinfiltration, and was reflected by reduced chlorophyll content. However, in these experiments, XopAP did not induce significant increase in ion leakage in the inoculated area (Teper et al., 2016). Results from this study indicate that XopAP acts as a virulence determinant in X. euvesicatoria, and contributes to the development of disease symptoms. A further study by Popov and colleagues (2018) revealed that XopAP was among the X. euvesicatoria 85-10 effectors that inhibited PAMP-triggered immunity, as assessed by inhibition of activation of a flg22-inducible reporter gene in Arabidopsis protoplasts (Popov et al., 2018). Expression of XopAP in an attenuated mutant of Pseudomonas syringae pv. tomato (DC3000 ΔCEL) increased its virulence on tomato. Also, the DC3000 ΔCEL strain carrying xopAP induced decreased callose deposition in Arabidopsis cell walls than the DC3000 ΔCEL strain (Popov et al., 2018). XopAP shares similarity with the Ralstonia solanacearum type III effector RipAL and both effectors possess a putative lipase domain (Peeters et al., 2013; Teper et al., 2016). R. solanacearum RipAL was shown to suppress salicylic acid-mediated defense responses and induce jasmonic acid production in N. benthamiana (Nakano & Mukaihara, 2018). Mutations in the putative catalytic residues within the lipase-like domain of RipAL abolished these activities (Nakano & Mukaihara, 2018).

Virulence and infection of X. oryzae pv. oryzicola (Xoc) increased in transgenic rice (Oryza sativa L.) plants overexpressing xopAP. XopAP inhibited the acidification of vacuoles by competing with vacuolar H⁺ -pyrophosphatase (V-PPase) for binding to PtdIns(3,5)P₂ , leading to stomatal opening. Transgenic rice overexpressing XopAP also showed inhibition of stomatal closure when challenged by Xoc infection and treatment with the PAMP flg22. Moreover, XopAP suppressed flg22-induced gene expression, reactive oxygen species burst and callose deposition in host plants, demonstrating that XopAP subverts PAMP-triggered immunity during Xoc infection. Taken together, these findings demonstrated that XopAP overcomes stomatal immunity in plants by binding to lipids (Liu et al., 2022).

Unknown. Subcellular localization analyses of the R. solanacearum homolog, RipAL, suggested that RipAL localizes to chloroplasts and targets chloroplast lipids in plant cells (Nakano & Mukaihara, 2018).

Unknown. XopAP contains a putative lipase domain (lipase class 3 family domain; conserved protein domain family PLN03037) in amino acid positions 236-322 (Teper et al., 2016).

XopAP was found to bind to phosphatidylinositol 3,5-bisphosphate (PtdIns(3,5)P₂ ), a membrane phospholipid that functions in pH control in lysosomes, membrane dynamics, and protein trafficking (Liu et al., 2022).

Yes (e.g., X. campestris, X. axonopodis, X. perforans, X. citri, X. alfalfae, X. prunicola, X. phaseoli, X. hortorum, X. arboricola, X. translucens, X. oryzae, X. hyacinthi, X. transluscens) (e.g Potnis et al., 2011; Jalan et al., 2013; Peng et al., 2016; Constantin et al., 2017).

Yes (Ralstonia solanacearum, plant-pathogenic Acidovorax species, Brenneria rubrifaciens, Robbsia andropogonis).

Constantin EC, Haegeman A, Van Vaerenbergh J, Baeyen S, Van Malderghem C, Maes M, Cottyn B (2017). Pathogenicity and virulence gene content of Xanthomonas strains infecting Araceae, formerly known as Xanthomonas axonopodis pv. dieffenbachiae. Plant Pathol. 66: 1539-1554. DOI: 10.1111/ppa.12694

Jalan N, Kumar D, Andrade MO, Yu F, Jones JB, Graham JH, White FF, Setubal JC, Wang N (2013). Comparative genomic and transcriptome analyses of pathotypes of Xanthomonas citri subsp. citri provide insights into mechanisms of bacterial virulence and host range. BMC Genomics 14: 551. DOI: 10.1186/1471-2164-14-551

Liu L, Li Y, Xu Z, Chen H, Zhang J, Manion B, Liu F, Zou L, Fu ZQ, Chen G (2022). The Xanthomonas type III effector XopAP prevents stomatal closure by interfering with vacuolar acidification. J. Integr. Plant Biol. 64: 1994-2008. DOI: 10.1111/jipb.13344

Nakano M, Mukaihara T (2018). Ralstonia solanacearum type III effector RipAL targets chloroplasts and induces jasmonic acid production to suppress salicylic acid-mediated responses in plants. Plant Cell Physiol. 59: 2576-2589. DOI: 10.1093/pcp/pcy177

Peeters N, Carrere S, Anisimova M, Plener L, Cazale AC, Genin S (2013). Repertoire, unified nomenclature and evolution of the type III effector gene set in the Ralstonia solanacearum species complex. BMC Genomics 14: 859. DOI: 10.1186/1471-2164-14-859

Peng, Z., Hu, Y., Xie, J., Potnis N, Akhunova A, Jones J, Liu Z, White FJ, Liu S (2016). Long read and single molecule DNA sequencing simplifies genome assembly and TAL effector gene analysis of Xanthomonas translucens. BMC Genomics 17: 21. DOI: 10.1186/s12864-015-2348-9

Popov G, Fraiture M, Brunner F, Sessa G (2018). 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

Potnis N, Krasileva K, Chow V, Almeida NF, Patil PB, Ryan RP, Sharlach M, Behlau F, Dow JM, Momol M, White FF, Preston JF, Vinatzer BA, Koebnik R, Setubal JC, Norman DJ, Staskawicz BJ, Jones JB (2011). Comparative genomics reveals diversity among xanthomonads infecting tomato and pepper. BMC Genomics 12: 146. DOI: 10.1186/1471-2164-12-146

Teper D, Burstein D, Salomon D, Gershovitz M, Pupko T, Sessa G (2016). Identification of novel Xanthomonas euvesicatoria type III effector proteins by a machine-learning approach. Mol. Plant Pathol. 17: 398-411. DOI: 10.1111/mpp.12288

Zhang Y, Teper D, Xu J, Wang N (2019). Stringent response regulators (p)ppGpp and DksA positively regulate virulence and host adaptation of Xanthomonas citri. Mol. Plant Pathol. 20:1550-1565. DOI: 10.1111/mpp.12865.

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