Hpa2 Required by HrpF To Translocate Xanthomonas oryzae Transcriptional Activator-Like Effectors into Rice for Pathogenicity

School of Agriculture and Biology, Shanghai Jiaotong University, 800 Dongchuan Road, Shanghai 200240, China.
Applied and Environmental Microbiology (Impact Factor: 3.67). 06/2011; 77(11):3809-18. DOI: 10.1128/AEM.02849-10
Source: PubMed


Xanthomonas oryzae pv. oryzicola, the causative agent of bacterial leaf streak, injects a plethora of effectors through the type III secretion
system (T3SS) into rice cells to cause disease. The T3SS, encoded by the hrp genes, is essential for the pathogen to elicit the hypersensitive response (HR) in nonhost tobacco and for pathogenicity
in host rice. Whether or not a putative lytic transglycosylase, Hpa2, interacts with a translocon protein, HrpF, to facilitate
bacterial pathogenicity remains unknown. Here we demonstrated that both the hpa2 and hrpF genes are required for the pathogenicity of X. oryzae pv. oryzicola strain RS105 in rice but not for HR induction in tobacco. The expression of hpa2 was positively regulated by HrpG and HrpD6 but not by HrpX. In vivo secretion and subcellular localization analyses confirmed that Hpa2 secretion is dependent on HpaB (a T3SS exit protein)
and that Hpa2 binds to the host cell membrane. Protein-protein assays demonstrated that Hpa2 interacts with HrpF. In planta translocation of AvrXa10 indicated that the mutation in hpa2 and hrpF inhibits the injection of the HpaB-dependent transcriptional activator-like (TAL) effector into rice. These findings suggest
that Hpa2 and HrpF form a complex to translocate T3S effectors into plant cells for pathogenesis in host rice.

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Available from: Gongyou Chen, Jul 28, 2014
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    • "Four-week-old rice seedlings were inoculated by infiltrating secondary leaves with a needleless syringe (Li et al. 2011b). Water-soaked symptoms or an HR were recorded 2 dpi. "
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    ABSTRACT: The closely related plant pathogens Xanthomonas oryzae pv. oryzicola and X. oryzae pv. oryzae cause bacterial leaf streak (BLS) and bacterial leaf blight (BLB), respectively, in rice. Unlike X. oryzae pv. oryzae, endogenous avirulence-resistance (avr-R) gene interactions have not been identified in the X. oryzae pv. oryzicola-rice pathosystem, though both X. oryzae pv. oryzicola and X. oryzae pv. oryzae possess transcriptional activator-like effectors (TALE), which are known to modulate R or S genes in rice. In this report, avrXa7, avrXa10, and avrXa27 from X. oryzae pv. oryzae were transferred into YNB0-17 and RS105, hypovirulent and hypervirulent strains, respectively, of X. oryzae pv. oryzicola. When YNB0-17 containing avrXa7, avrXa10, or avrXa27 was inoculated to rice, hypersensitive responses (HR) were elicited in rice cultivars containing the R genes Xa7, Xa10, and Xa27, respectively. By contrast, RS105 expressing avrXa27 elicited an HR in a rice cultivar containing Xa27 but the expression of avrXa7 and avrXa10 in RS105 did not result in HR in rice cultivars containing Xa7 and Xa10, correspondingly. Southern blot analysis demonstrated that YNB0-17 possesses only approximately nine putative tale genes, whereas the hypervirulent RS105 contains at least 20. Although YNB0-17 contains an intact type III secretion system (T3SS), its genome is lacking the T3SS effector genes avrRxo1 and xopO, which are present in RS105. The introduction of avrRxo1 and xopO into YNB0-17 did not suppress avrXa7- or avrXa10-triggered immunity in rice. However, the transference of individual tale genes from RS105 into YNB0-17 led to the identification of tal6 and tal11a that suppressed avrXa7-Xa7-mediated defense. Thus, YNB0-17 may be a useful recipient for discovering such suppressors. This is the first report that co-evolutionally generated tale genes in X. oryzae pv. oryzicola suppress gene-for-gene defense against BLB, which may explain the lack of BLS-resistant cultivars.
    Molecular Plant-Microbe Interactions 09/2014; 27(9):983-95. DOI:10.1094/MPMI-09-13-0279-R · 3.94 Impact Factor
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    • "RDhrpG hrpG deletion mutant of strain RS105, Rif r Jiang et al. (2009a) RDhrpX hrpX deletion mutant of strain RS105; Rif r Jiang et al. (2009a) RDhpaB hpaB deletion mutant of strain RS105, Rif r Li et al. (2011b) RDhrcV hrcV deletion mutant of strain RS105; Rif r This laboratory RDhrpD5 hrpD5 deletion mutant of strain RS105, Rif r Li et al. (2011a) RDhrpD6 hrpD6 deletion mutant of strain RS105, Rif r Li et al. (2011a) RDhrpE hrpE deletion mutant of strain RS105, Rif r Li et al. (2011a) "
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    ABSTRACT: Xanthomonas oryzae pv. oryzicola (Xoc) is the causal agent of bacterial leaf streak, a devastating disease in rice. Xoc uses a type III secretion (T3S) system, which is encoded by the hrp-hrc-hpa (hypersensitive response and pathogenicity, hrp-conserved and hrp-associated) genes, to inject repertoires of T3S effectors (T3Es) into plant cells. Many of the hrp-hrc-hpa genes have roles in pathogenesis, but the role of hrpE3, which shows homology to hpaE in X. campestris pv. vesicatoria (Xcv), is poorly understood. In this study, hrpE3 was shown to be transcribed independent of the hrpD operon, and its expression was dependent on a promoter within hpaB. The expression of hrpE3 was positively regulated by HrpG and HrpX, a finding probably caused by an imperfect plant-inducible promoter (PIP) box (TTCGT-N16 -TTCGA) in the hrpE3 promoter. The secretion of HrpE3 was dependent on T3S, and subcellular localization of HrpE3 was cytoplasmic and nuclear in plant cells. A mutation in hrpE3 reduced the virulence of Xoc by decreasing disease lesion length and bacterial growth in planta. Full virulence was restored to the mutant when Xoc hrpE3, but not Xcv hpaE, was expressed in trans. The differences in transcription, secretion via the T3S system and bacterial virulence in plants were attributed to N-terminal amino acid differences between Xoc HrpE3 and Xcv HpaE. Collectively, the results demonstrate that hrpE3 encodes a T3E protein which is delivered into the plant cell through the T3S system, localizes to the cytoplasm and nucleus, and is required for full virulence in rice.
    Molecular Plant Pathology 05/2013; 14(7). DOI:10.1111/mpp.12039 · 4.72 Impact Factor
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    • "Bacterial strains and plasmids used in this study are listed in Table 1. The Xoc RS105 wild-type and mutant strains were grown in NB medium (beef extract, 3 g/L; yeast extract, 1 g/L; tryptone, 5 g/L; sucrose, 10 g/L ), NYG medium or in XOM3, T3SS-inducing minimal medium [56] at 28°C. Antibiotics were used at the following concentrations: ampicillin, 100 µg/ml; kanamycin, 50 µg/ml; rifampin (Rif), 25 µg/ml. "
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    ABSTRACT: Bacterial leaf streak caused by Xanthomonas oryzae pv. oryzicola (Xoc) is one of the most important diseases in rice. However, little is known about the pathogenicity mechanisms of Xoc. Here we have investigated the function of three HD-GYP domain regulatory proteins in biofilm formation, the synthesis of virulence factors and virulence of Xoc. Deletion of rpfG resulted in altered production of extracellular polysaccharides (EPS), abolished virulence on rice and enhanced biofilm formation, but had little effect on the secretion of proteases and motility. In contrast, mutational analysis showed that the other two HD-GYP domain proteins had no effect on virulence factor synthesis and tested phenotypes. Mutation of rpfG led to up-regulation of the type III secretion system and altered expression of three putative glycosyltransferase genes gumD, pgaC and xagB, which are part of operons directing the synthesis of different extracellular polysaccharides. The pgaABCD and xagABCD operons were greatly up-regulated in the Xoc ΔrpfG mutant, whereas the expression of the gum genes was unaltered or slightly enhanced. The elevated biofilm formation of the Xoc ΔrpfG mutant was dramatically reduced upon deletion of gumD, xagA and xagB, but not when pgaA and pgaC were deleted. Interestingly, only the ΔgumD mutant, among these single gene mutants, exhibits multiple phenotype alterations including reduced biofilm and EPS production and attenuated virulence on rice. These data indicate that RpfG is a global regulator that controls biofilm formation, EPS production and bacterial virulence in Xoc and that both gumD- and xagB-dependent EPS contribute to biofilm formation under different conditions.
    PLoS ONE 03/2013; 8(3):e59428. DOI:10.1371/journal.pone.0059428 · 3.23 Impact Factor
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