Research ArticlePlant biology

The plant cell wall integrity maintenance and immune signaling systems cooperate to control stress responses in Arabidopsis thaliana

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Science Signaling  26 Jun 2018:
Vol. 11, Issue 536, eaao3070
DOI: 10.1126/scisignal.aao3070
  • Fig. 1 Different types of CWD induce similar osmosensitive responses.

    Quantification of (A) JA and (B) SA, expressed as microgram per gram dry weight (gDW), in Col-0, bak1-5, and ixr1-1 seedlings that had been treated with DMSO (mock), DMSO and sorbitol (S), ISX, or ISX and sorbitol (ISX + S). (C) JA and (D) SA quantification in Col-0, bak1-5, and ixr1-1 seedlings treated with boiled (inactive) Driselase (bDri), bDri and sorbitol (bDri + S), Driselase (Dri), or Driselase and sorbitol (Dri + S). (E) Relative expression of PDF1.2 as determined by quantitative reverse transcription polymerase chain reaction (qRT-PCR) in Col-0 and bak1-5 seedlings treated with bDri or Dri compared to untreated (NT) seedlings. (F) Relative expression of TCH4 in Col-0 seedlings treated as indicated. (G) JA and (H) SA quantification in Col-0 seedlings treated with boiled pectinase (bP), boiled pectinase and sorbitol (bP + S), pectinase (P), or pectinase and sorbitol (P + S). (I) JA and (J) SA quantification in Col-0 seedlings treated with the indicated combinations of boiled (b) or active preparations of cellulase (C), pectinase (P), xylanase (X), and sorbitol (S). All values represent means with error bars indicating SD. n = 4 (A to D) and n = 3 (E to J). Letters a to d (A to J) indicate statistically significant differences according to one-way analysis of variance (ANOVA) and Tukey’s HSD (honestly significantly different) test (α = 0.05) between treatments for each genotype. Asterisks (A to D) indicate statistically significant differences to the wild type (Student’s t test, **P < 0.01; ***P < 0.001; ns, not significant).

  • Fig. 2 Phenotypic clustering identifies groups of genes involved in CWD responses.

    Quantification of (A) root tip lignification, (B) JA, and (C) SA in the indicated mutant seedlings after treatment with ISX. Values represent means with error bars indicating SD and are expressed relative to the appropriate wild-type control (Col-0 or Ws-2, depending on the genetic background of the mutant strain) from a representative experiment (dashed line). n ≥ 10 (A) and n = 4 (B and C); asterisks indicate statistically significant differences between the mutant and wild type (Student’s t test, *P < 0.05). Mutant lines are organized in functional groups (RLKs, CrRLK1Ls, AHKs, and Ion channels), and individual genotypes are described in detail in table S2. (D) Hierarchical clustering of mutant phenotypes assigning functions in CWI maintenance to candidate genes based on their responses to ISX. Mutant phenotype data from (A) to (C) and fig. S9F [root growth inhibition (RGI)] were normalized to wild-type controls and log2-transformed before average linkage clustering. Blue color indicates reduced ISX responses, and red color indicates increased ISX responses compared to wild type.

  • Fig. 3 THE1 functions upstream of MCA1 and FEI2 and promotes responses to different types of CWD.

    Quantification of (A) root tip lignification, (B) JA, and (C) SA after ISX treatment in wild-type (Col-0) and mutant seedlings carrying the indicated loss-of-function mutations. Quantification of (D) root tip lignification, (E) JA, and (F) SA after ISX treatment in Col-0 and mutant seedlings carrying the gain-of-function allele the1-4 or the1-4 in combination with the loss-of-function allele mca1 or fei2. Quantification of (G) JA and (H) SA in Col-0, the1-1 (loss-of-function), and the1-4 (gain-of-function) seedlings treated with boiled Driselase (bDri) or Driselase (Dri). n ≥ 17 (lignin) and n = 4 (JA and SA). Letters a to e (A to F) indicate statistically significant differences between genotypes according to one-way ANOVA and Tukey’s HSD test (α = 0.05). Asterisks (G and H) indicate statistically significant differences compared to the Col-0 control (Student’s t test, *P < 0.05; **P < 0.01; ***P < 0.001).

  • Fig. 4 ISX induces PROPEP expression, and AtPep1 and AtPep3 repress responses to CWD.

    (A) Relative PROPEP1, PROPEP2, PROPEP3, and PROPEP4 expression determined by qRT-PCR in Col-0 seedlings treated with DMSO (mock) or ISX for 1 hour. (B) PROPEP1 and PROPEP3 expression in Col-0 seedlings at the indicated time points. (C) PROPEP1 and PROPEP3 expression in Col-0 and the1-1 seedlings after 1 hour of mock or ISX treatment. Quantification of (D) JA and (E) SA in Col-0 seedlings after cotreatment with either mock conditions or ISX plus 0, 1, 10, or 100 nM AtPep1. Quantification of (F) JA and (G) SA in Col-0, pepr1, pepr2, and pepr1 pepr2 seedlings after cotreatment with either mock conditions or ISX plus 10 nM AtPep1. (H) Root tip lignification in Col-0, pepr1, pepr2, and pepr1 pepr2 seedlings after cotreatment with either mock conditions or ISX plus 10 nM AtPep1 was visualized by phloroglucinol staining. Scale bar, 200 μm. Quantification of (I) JA and (J) SA in Col-0 seedlings after cotreatment with either mock conditions or ISX plus 0 or 10 nM AtPep3. All values represent means with error bars indicating SD. n = 3 (A to C, I, and J), n = 4 (D to G), and n ≥ 5 (H). Asterisks (A to C) indicate statistically significant differences compared to mock-treated controls (Student’s t test, *P < 0.05). Letters a to e (D to J) indicate statistically significant differences between treatments of each genotype according to one-way ANOVA and Tukey’s HSD test (α = 0.05). (K) Seedlings expressing PROPEP3-Venus were mock- or ISX-treated for 24 hours. Proteins in the growth medium were immunoprecipitated, separated by SDS–polyacrylamide gel electrophoresis (PAGE), and stained with silver nitrate. The expected mass of PROPEP3-Venus is 37 kDa, and the expected mass of Venus alone is 27 kDa. n = 2. (L) Numbers of unique PROPEP3-Venus peptides after trypsin digest identified by LC-MS/MS from silver-stained bands at around 25 and 37 kDa. FDR, false discovery rate.

  • Fig. 5 Model of stress response integration through CWI and PTI signaling.

    (A) Responses to CWD caused by inhibition of cellulose biosynthesis (ISX) or enzymatic cell wall degradation (Driselase) in Arabidopsis depend on the RLK THE1. THE1 acts upstream of the ion channel MCA1 and the RLK FEI2 to stimulate the CWI maintenance system. Independently, through an unknown mechanism, ISX treatment induces the expression of PROPEP1 and PROPEP3 (PROPEP1/3) and secretion of PROPEP3. Processing of PROPEP1 and PROPEP3 generates the host defense peptides AtPep1 and AtPep3 (AtPep1/3), which are DAMPs that are also induced by pathogen elicitors during PTI. AtPep1/3 repress ISX-induced hormone accumulation through the AtPep receptors PEPR1 and PEPR2, suggesting an AtPep-dependent negative feedback mechanism. AtPep signaling also induces a positive feedback loop that enhances PTI. (B) If PTI-mediated induction of AtPeps or AtPep signaling is impaired, suppression of CWI signaling is alleviated, and the CWI maintenance pathway contributes to stress response induction to a greater extent than does PTI.

Supplementary Materials

  • www.sciencesignaling.org/cgi/content/full/11/536/eaao3070/DC1

    Fig. S1. Sorbitol modifies root responses to ISX and Driselase.

    Fig. S2. Sorbitol modifies responses to ISX and Driselase in cotyledons.

    Fig. S3. CWD-induced cell death and lignin and callose production are sensitive to turgor manipulation.

    Fig. S4. Sorbitol does not reduce OG- and flg22-induced defense responses.

    Fig. S5. Individual cell wall–degrading enzymes have specific effects on JA and SA accumulation.

    Fig. S6. Supernatants from damaged seedlings do not induce pronounced JA and SA accumulation.

    Fig. S7. ISX-induced CWD responses in osmosensing and mechanoperception mutants.

    Fig. S8. Isolation of the wak2-12 allele.

    Fig. S9. JA and SA accumulation, root growth, and ISX resistance phenotypes in mock-treated mutant seedlings.

    Fig. S10. THE1 is not required for flg22-induced SA accumulation.

    Fig. S11. PROPEP1-7 expression in ISX-treated seedlings and effects of AtPep1 and AtPep3 treatments on lignin production.

    Table S1. GO enrichment analysis of genes with ISX-dependent expression changes.

    Table S2. Arabidopsis genotypes used in this study.

    Table S3. Primers used in this study.

    Data file S1. Transcriptomics data.

    Data file S2. Peptide mass fingerprinting data.

    References (7891)

  • Supplementary Materials for:

    The plant cell wall integrity maintenance and immune signaling systems cooperate to control stress responses in Arabidopsis thaliana

    Timo Engelsdorf, Nora Gigli-Bisceglia, Manikandan Veerabagu, Joseph F. McKenna, Lauri Vaahtera, Frauke Augstein, Dieuwertje Van der Does, Cyril Zipfel, Thorsten Hamann*

    *Corresponding author. Email: thorsten.hamann{at}ntnu.no

    This PDF file includes:

    • Fig. S1. Sorbitol modifies root responses to ISX and Driselase.
    • Fig. S2. Sorbitol modifies responses to ISX and Driselase in cotyledons.
    • Fig. S3. CWD-induced cell death and lignin and callose deposition are sensitive to turgor manipulation.
    • Fig. S4. Sorbitol does not reduce OG- and flg22-induced defense responses.
    • Fig. S5. Individual cell wall–degrading enzymes have specific effects on JA and SA accumulation.
    • Fig. S6. Supernatants from damaged seedlings do not induce pronounced JA and SA accumulation.
    • Fig. S7. ISX-induced CWD responses in osmosensing and mechanoperception mutants.
    • Fig. S8. Isolation of the wak2-12 allele.
    • Fig. S9. JA and SA accumulation, root growth, and ISX resistance phenotypes in mock-treated mutant seedlings.
    • Fig. S10. THE1 is not required for flg22-induced SA accumulation.
    • Fig. S11. PROPEP1-7 expression in ISX-treated seedlings and effects of AtPep1 and AtPep3 treatments on lignin production.
    • Table S1. GO enrichment analysis of genes with ISX-dependent expression changes.
    • Table S2. Arabidopsis genotypes used in this study.
    • Table S3. Primers used in this study.
    • Legends for data files S1 and S2
    • References (7891)

    [Download PDF]

    Other Supplementary Material for this manuscript includes the following:

    • Data file S1 (Microsft Excel format). Transcriptomics data.
    • Data file S2 (Microsft Excel format). Peptide mass fingerprinting data.

    [Download Data files S1 and S2]


    © 2018 American Association for the Advancement of Science

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