Extended Data Fig. 10: TCP21 is required for the HR induction by Tsw and NSs in N. benthamiana plant and required for Tsw-mediated resistance in pepper against TSWV infection.
From: NLR surveillance of pathogen interference with hormone receptors induces immunity
a, Effect of CaTCP21-1 on the HR cell death induced by co-expression of Tsw and NSs. CaTCP21-1 (i), Tsw (ii), Tsw+CaTCP21-1 (iii), Tsw+NSs+EV (iv) or Tsw+NSs+CaTCP21-1 (v) was/were (co)expressed in N. benthamiana leaves via agroinfiltration. The HR was monitored from 0-8 dpi and photographed at 5 dpi. b, Effect of CaTCP3 and CaTCP17 on the HR cell death induced by co-expression of Tsw and NSs. Data are represented as mean values ± s. e. m.; n = 6 biologically independent samples. c, Effect of CaTCP21-1 on the HR cell death induced by co-expression of Sw-5b NLR and NSm effector. HR phenotype was photographed at 5 dpi. The ion leakage of infiltrated leaves was measured at 2 dpi and shown in the right. Data are represented as mean values ± s. e. m.; n = 6 biologically independent samples. d, Schematic diagram of TRV-mediated gene silencing of NbTCP21 in N. benthamiana plant. TRV carrying a 300 bp cDNA fragment each of NbTCP21-1 and NbTCP21-2 (TRV-NbTCP21-1/2) was used to infect N. benthamiana plant. After 20 days post TRV treatment, the upper new leaves that silenced with TCP21 were co-expressed with Tsw and NSsY30A. The onset of HR was monitored from 0-8 dpi. e, The phenotype of N. benthamiana plant treated with TRV-GUS and TRV-NbTCP21-1/2 at 20 dpi. f, Relative RNA expression of NbTCP21-1 and NbTCP21-2 in N. benthamiana plant treated with TRV-GUS and TRV-NbTCP21-1/2. Data are presented as mean values ± s.e.m.; n = 3 independent plant samples. g, BiFC analysis of the interaction between NSs and Tsw-PRL in TRV-GUS treated control plant leaves or NbTCP21-1/2 silenced N. benthamiana plant leaves. The BiFC fluorescence signals were examined and photographed at 39 h post agroinfiltration. h, Silencing of NbTCP21-1/2 did not affect the HR induced by Sw-5b NLR and NSm effector. Sw-5b and NSm were coexpressed by agro-infiltration in N. benthamiana plant treated with TRV-GUS or TRV-NbTCP21-1/2. HR phenotype of infiltrated leaves was monitored and photographed at 5 dpi. The ion leakage of infiltrated leaves was measured at 2 dpi and shown in the right. Data are presented as mean values ± s.e.m.; n = 6 biologically independent samples. i, Phenotype of pepper “PI152225” silenced for CaTCP21-1/3/4/7 and Tsw. TRV carrying a 300 bp cDNA fragment of GUS (TRV-GUS), Tsw (TRV-Tsw) or the fusion of 200 bp cDNA fragment each of CaTCP21-1, 3, 4 and 7 (TRV-CaTCP21-1/3/4/7) was used to infect pepper “PI152225” carrying Tsw. The phenotype of pepper treated with various TRV constructs were photographed at 30 dpi. TRV-CaPDS served as a positive control for silencing (right). j-l, Relative expression levels of CaTCP21-1, -3, -4, -7, Tsw and CaPDS genes in pepper treated with TRV-GUS, TRV-CaTCP21-1/3/4/7 (j), TRV-Tsw (k) or TRV-CaPDS (l). Data are represented as mean values ± s. e. m.; n = 3 biologically independent samples. Data in (b, c) were analysed by one-way ANOVA with Tukey’s test; lowercase letters a and b represent statistically different groups (p < 0.05). Data in (f, h, j-l) were analysed by two-sided Student’s t-test. P values in (b, c, j) are shown in the Source Data; P values in (f, h, k, i) are shown in the figure. In box plots of (b, c, h), the center line represents the median, box edges delimit lower and upper quartiles and whiskers show the highest and lowest data points.
