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--- bacteria:t3e:xops [2025/07/04 23:46] – jfpothier
+++ bacteria:t3e:xops [2025/07/24 23:21] (current) – jfpothier
@@ Line 21: / Line 21: @@
 Type III secretion (T3S) assays of XopS-AvrBs3Δ2 fusion proteins indicated that XopS is secreted and translocated into the plant cells through T3S, inducing HR in Bs3 resistant pepper leaves. Further supporting the identify of XopS as a T3E, it was shown that deletion mutants (Δ//xopS//) cause a considerable attenuation of disease symptoms in pepper (Schulze //et al.//, 2012).
-For expression in //X. campestris// pv. //vesicatoria// (//Xcv//), //xopS// was amplified from strain 85-10 and cloned into the Golden Gate‐compatible expression vector pBRM (Schulze //et al//., 2012). To generate //avrBs3//Δ2 fusions, the promoters and 5′ coding sequences of //xopS// was amplified by PCR from genomic DNA of //Xcv// 85-10, cloned into pENTR/D‐TOPO and recombined into pL6GW356 (Schulze //et al//., 2012). To generate deletions of //xopS//, 0.6–1kb fragments upstream and downstream of the respective gene were amplified from genomic DNA of //Xcv// 85-10 by PCR using oligonucleotides harboring appropriate restriction sites. Fragments were cloned into the suicide vectors pOK1 (Schulze //et al//., 2012).
+For expression in //X. campestris// pv. //vesicatoria// (//Xcv//), //xopS// was amplified from strain 85-10 and cloned into the Golden Gate‐compatible expression vector pBRM (Schulze //et al//., 2012). To generate //avrBs3//Δ2 fusions, the promoters and 5' coding sequences of //xopS// was amplified by PCR from genomic DNA of //Xcv// 85-10, cloned into pENTR/D‐TOPO and recombined into pL6GW356 (Schulze //et al//., 2012). To generate deletions of //xopS//, 0.6–1kb fragments upstream and downstream of the respective gene were amplified from genomic DNA of //Xcv// 85-10 by PCR using oligonucleotides harboring appropriate restriction sites. Fragments were cloned into the suicide vectors pOK1 (Schulze //et al//., 2012).
 === Regulation ===
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 To study the contribution of the T3Es to bacterial virulence, the effector gene was individually deleted in //Xcv// strain 85‐10, and the mutant was inoculated into leaves of susceptible ECW pepper plants. In addition, induction of the HR in pepper ECW‐10R was analyzed, which is based on the recognition of the T3E AvrBs1 by the Bs1 resistance gene (Schulze //et al//., 2012). Schulze et al. 2012 studied XopS along with XopB in their study. Deletion of //xopB// or //xopS// led to significantly reduced disease symptoms, whereas the HR induction was not impaired. The mutant phenotypes of 85-10Δ//xopB// and 85-10Δ//xopS// were complemented by ectopic expression of the respective effector gene, suggesting that reduced virulence was not caused by polar effects of the deletions on downstream genes. Although the growth of both individual effector mutants in ECW plants did not differ significantly from that of the wild‐type strain), multiplication of an 85‐10Δ//xopB//Δ//xopS// double mutant was reduced significantly, suggesting that XopB and XopS fulfill redundant functions. To identify additional virulence phenotypes, as well as defense reactions, mediated by the analyzed T3Es, leaves of pepper ECW, //Nicotiana benthamiana// and //N. tabacum// were inoculated with //Agrobacterium// strains mediating the in planta expression of the effector genes fused to GFP. These experiments confirmed that XopS (similar to XopB, XopG, XopM) trigger cell death in different //Solanaceae// (Schulze //et al//., 2012). XopS is involved in the severity of disease symptoms, the promotion of bacterial growth and the suppression of PTI (Schulze //et al//., 2012).
-XopS<sub>//Xcv//85-10</sub> inhibited proteasomal degradation of WRKY40, a transcriptional regulator of defense gene expression. Virus-induced gene silencing of WRKY40 in pepper enhanced plant tolerance to //Xcv// infection, indicating that WRKY40 represses immunity. Stabilization of WRKY40 by XopS reduced the expression of its targets, which included salicylic acid-responsive genes and the jasmonic acid signaling repressor JAZ8. //Xcv// bacteria lacking XopS displayed significantly reduced virulence when surface inoculated onto susceptible pepper leaves. XopS delivery by //Xcv//, as well as ectopic expression of XopS in //Arabidopsis thaliana// or //Nicotiana benthamiana//, prevented stomatal closure in response to bacteria and biotic elicitors. Silencing WRKY40 in pepper or //N. benthamiana// abolished XopS’s ability to prevent stomatal closure. These findings suggests that XopS interferes with both preinvasion and apoplastic defense by manipulating WRKY40 stability and downstream gene expression, eventually altering phytohormone crosstalk to promote pathogen proliferation (Raffeiner //et al.//, 2022).
+XopS<sub>//Xcv//85-10</sub> inhibited proteasomal degradation of WRKY40, a transcriptional regulator of defense gene expression. Virus-induced gene silencing of WRKY40 in pepper enhanced plant tolerance to //Xcv// infection, indicating that WRKY40 represses immunity. Stabilization of WRKY40 by XopS reduced the expression of its targets, which included salicylic acid-responsive genes and the jasmonic acid signaling repressor JAZ8. //Xcv// bacteria lacking XopS displayed significantly reduced virulence when surface inoculated onto susceptible pepper leaves. XopS delivery by //Xcv//, as well as ectopic expression of XopS in //Arabidopsis thaliana// or //Nicotiana benthamiana//, prevented stomatal closure in response to bacteria and biotic elicitors. Silencing WRKY40 in pepper or //N. benthamiana// abolished XopS's ability to prevent stomatal closure. These findings suggests that XopS interferes with both preinvasion and apoplastic defense by manipulating WRKY40 stability and downstream gene expression, eventually altering phytohormone crosstalk to promote pathogen proliferation (Raffeiner //et al.//, 2022).
 === Localization ===

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