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A candidate RxLR effector from Plasmopara viticola can elicit immune responses in Nicotiana benthamiana

View Article: PubMed Central - PubMed

ABSTRACT

Background: Diverse plant pathogens deliver effectors into plant cells to alter host processes. Oomycete pathogen encodes a large number of putative RxLR effectors which are likely to play a role in manipulating plant defense responses. The secretome of Plasmopara viticola (downy mildew of grapevine) contains at least 162 candidate RxLR effectors discovered in our recent studies, but their roles in infection and pathogenicity remain to be determined. Here, we characterize in depth one of the putative RxLR effectors, PvRxLR16, which has been reported to induce cell death in Nicotiana benthamiana in our previous study.

Results: The nuclear localization, W/Y/L motifs, and a putative N-glycosylation site in C-terminal of PvRxLR16 were essential for cell death-inducing activity. Suppressor of G-two allele of Skp1 (SGT1), heat shock protein 90 (HSP90) and required for Mla12 resistance (RAR1), but not somatic embryogenesis receptor-like kinase (SERK3), were required for the cell death response triggered by PvRxLR16 in N. benthamiana. Some mitogen-activated protein kinases and transcription factors were also involved in the perception of PvRxLR16 by N. benthamiana. PvRxLR16 could also significantly enhance plant resistance to Phytophthora capsici and the nuclear localization was required for this ability. However, some other PvRxLR effectors could suppress defense responses and disease resistance induced by PvRxLR16, suggesting that it may not trigger host cell death or immune responses during physiological infection under natural conditions.

Conclusion: These data demonstrate that PvRxLR16 may be recognized by endogenous proteins in nucleus to trigger immune responses in N. benthamiana, which in turn can be suppressed by other PvRxLR effectors.

Electronic supplementary material: The online version of this article (doi:10.1186/s12870-017-1016-4) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus

MAPK cascades were required for PvRxLR16-induced cell death in N. benthamiana. a PvRxLR16 was transiently expressed in N. benthamiana leaves silenced for indicated MAPK cascades genes. GFP and INF1 are control proteins. Typical symptoms were photographed after 5 d after agroinfiltration. b Quantification of cell death by measuring electrolyte leakage. Averages and standard errors were calculated from three independent experimental treatments (**, P < 0.01, Dunnett’s test). c Transcript levels of indicated genes in silenced N. benthamiana measured by quantitative RT-PCR. Error bars represent standard errors from three biological replicates (**, P < 0.01, Dunnett’s test)
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Fig6: MAPK cascades were required for PvRxLR16-induced cell death in N. benthamiana. a PvRxLR16 was transiently expressed in N. benthamiana leaves silenced for indicated MAPK cascades genes. GFP and INF1 are control proteins. Typical symptoms were photographed after 5 d after agroinfiltration. b Quantification of cell death by measuring electrolyte leakage. Averages and standard errors were calculated from three independent experimental treatments (**, P < 0.01, Dunnett’s test). c Transcript levels of indicated genes in silenced N. benthamiana measured by quantitative RT-PCR. Error bars represent standard errors from three biological replicates (**, P < 0.01, Dunnett’s test)

Mentions: MAPK cascades play a remarkably important role in both PTI and ETI [53]. A series of kinases and transcription factors have been characterized that play an essential role in plant immunity and cell death induction during interactions between plants and pathogens, including mitogen-activated protein kinase kinase kinase (MAPKKKα), MAP kinase kinase 2 (MEK2), salicylic acid-induced protein kinase (SIPK), MAP kinase kinase 1 (MEK1), NTF6, wound-induced protein kinase (WIPK), WRYK1 and WRKY2 [54–56]. To investigate the possible roles of these proteins in PvRxLR16-induced cell death, each of them was silenced in N. benthamiana via VIGS and challenged with PvRxLR16 effector. INF1 was chosen as a positive control which triggers cell death independently of these kinases and transcription factors. Cell death was compromised in plants silenced for all genes with one exception (WIPK) (Fig. 6a and b). Figure 6c showed the transcript abundances of purpose genes in the silenced N. benthamiana, validating successful silencing. These results suggest that most mitogen-activated protein kinases and transcription factors tested in this study are involved in the perception of PvRxLR16 by N. benthamiana.Fig. 6


A candidate RxLR effector from Plasmopara viticola can elicit immune responses in Nicotiana benthamiana
MAPK cascades were required for PvRxLR16-induced cell death in N. benthamiana. a PvRxLR16 was transiently expressed in N. benthamiana leaves silenced for indicated MAPK cascades genes. GFP and INF1 are control proteins. Typical symptoms were photographed after 5 d after agroinfiltration. b Quantification of cell death by measuring electrolyte leakage. Averages and standard errors were calculated from three independent experimental treatments (**, P < 0.01, Dunnett’s test). c Transcript levels of indicated genes in silenced N. benthamiana measured by quantitative RT-PCR. Error bars represent standard errors from three biological replicates (**, P < 0.01, Dunnett’s test)
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Related In: Results  -  Collection

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Fig6: MAPK cascades were required for PvRxLR16-induced cell death in N. benthamiana. a PvRxLR16 was transiently expressed in N. benthamiana leaves silenced for indicated MAPK cascades genes. GFP and INF1 are control proteins. Typical symptoms were photographed after 5 d after agroinfiltration. b Quantification of cell death by measuring electrolyte leakage. Averages and standard errors were calculated from three independent experimental treatments (**, P < 0.01, Dunnett’s test). c Transcript levels of indicated genes in silenced N. benthamiana measured by quantitative RT-PCR. Error bars represent standard errors from three biological replicates (**, P < 0.01, Dunnett’s test)
Mentions: MAPK cascades play a remarkably important role in both PTI and ETI [53]. A series of kinases and transcription factors have been characterized that play an essential role in plant immunity and cell death induction during interactions between plants and pathogens, including mitogen-activated protein kinase kinase kinase (MAPKKKα), MAP kinase kinase 2 (MEK2), salicylic acid-induced protein kinase (SIPK), MAP kinase kinase 1 (MEK1), NTF6, wound-induced protein kinase (WIPK), WRYK1 and WRKY2 [54–56]. To investigate the possible roles of these proteins in PvRxLR16-induced cell death, each of them was silenced in N. benthamiana via VIGS and challenged with PvRxLR16 effector. INF1 was chosen as a positive control which triggers cell death independently of these kinases and transcription factors. Cell death was compromised in plants silenced for all genes with one exception (WIPK) (Fig. 6a and b). Figure 6c showed the transcript abundances of purpose genes in the silenced N. benthamiana, validating successful silencing. These results suggest that most mitogen-activated protein kinases and transcription factors tested in this study are involved in the perception of PvRxLR16 by N. benthamiana.Fig. 6

View Article: PubMed Central - PubMed

ABSTRACT

Background: Diverse plant pathogens deliver effectors into plant cells to alter host processes. Oomycete pathogen encodes a large number of putative RxLR effectors which are likely to play a role in manipulating plant defense responses. The secretome of Plasmopara viticola (downy mildew of grapevine) contains at least 162 candidate RxLR effectors discovered in our recent studies, but their roles in infection and pathogenicity remain to be determined. Here, we characterize in depth one of the putative RxLR effectors, PvRxLR16, which has been reported to induce cell death in Nicotiana benthamiana in our previous study.

Results: The nuclear localization, W/Y/L motifs, and a putative N-glycosylation site in C-terminal of PvRxLR16 were essential for cell death-inducing activity. Suppressor of G-two allele of Skp1 (SGT1), heat shock protein 90 (HSP90) and required for Mla12 resistance (RAR1), but not somatic embryogenesis receptor-like kinase (SERK3), were required for the cell death response triggered by PvRxLR16 in N. benthamiana. Some mitogen-activated protein kinases and transcription factors were also involved in the perception of PvRxLR16 by N. benthamiana. PvRxLR16 could also significantly enhance plant resistance to Phytophthora capsici and the nuclear localization was required for this ability. However, some other PvRxLR effectors could suppress defense responses and disease resistance induced by PvRxLR16, suggesting that it may not trigger host cell death or immune responses during physiological infection under natural conditions.

Conclusion: These data demonstrate that PvRxLR16 may be recognized by endogenous proteins in nucleus to trigger immune responses in N. benthamiana, which in turn can be suppressed by other PvRxLR effectors.

Electronic supplementary material: The online version of this article (doi:10.1186/s12870-017-1016-4) contains supplementary material, which is available to authorized users.

No MeSH data available.


Related in: MedlinePlus