Linker histone H1 modulates defense priming and immunity in plants
Item Type Article
Authors Sheikh, Arsheed Hussain;Nawaz, Kashif;Tabassum,
Naheed;Trapp, Marilia Almeida;Mariappan, Kiruthiga;Alhoraibi, Hanna;Rayapuram, Naganand;Aranda, Manuel;Groth, Martin;Hirt, Heribert
Citation Sheikh, A. H., Nawaz, K., Tabassum, N., Almeida-Trapp, M., Mariappan, K. G., Alhoraibi, H., Rayapuram, N., Aranda, M., Groth, M., & Hirt, H. (2023). Linker histone H1 modulates defense priming and immunity in plants. Nucleic Acids Research. https://
doi.org/10.1093/nar/gkad106 Eprint version Publisher's Version/PDF
DOI 10.1093/nar/gkad106
Publisher Oxford University Press (OUP) Journal Nucleic Acids Research
Rights Archived with thanks to Nucleic Acids Research under a Creative Commons license, details at: https://creativecommons.org/
licenses/by-nc/4.0/
Download date 2024-01-25 22:16:55
Item License https://creativecommons.org/licenses/by-nc/4.0/
Link to Item http://hdl.handle.net/10754/689139
Supplementary Figures
Supplementary Figure S1. Innate immunity (PTI) levels in 3h1
(A) Reactive oxygen species (ROS) levels quantified in WT and 3h1 leaf discs triggered over 40 min by chitin treatment.
(B) MAPK activation shown by anti-pTEpY western blot indicates the phosphorylation of MPK6, MPK3 and MPK4 in 2-week-old WT and 3h1 seedlings after 1 μM flg22 treatment for 30 min. CBB stain of Rubisco serves as a loading control.
(C, D) Expression of innate immunity genes FRK1 and MYB51 after 6h of Pst DC3000 infection.
(E) Jasmonic acid (JA) quantification in WT and 3h1 plants is shown as ng/g of fresh weight.
The data are shown as means ± SEMs from three replicates. Asterisk indicates a significant difference with P < 0.05.
Supplementary Figure S2. Transcriptome of 3h1 plants
(A) Principal component analysis (PCA) plot of the RNA-seq of WT and 3h1 plants treated with Pst DC3000 infection at 6 and 24 h of infection.
(B) Venn diagrams showing the number of upregulated and downregulated genes in 3h1 as compare to WT after Pst DC3000 infection.
(C) Principal component analysis (PCA) plot of the transcriptome of flg22 pretreated WT and 3h1 plants challenged with Pst DC3000 infection at 6 and 24 h of infection.
(D) Venn diagrams showing the number of upregulated and downregulated genes in flg22 treated 3h1 as compared to flg22 treated WT and then challenged by Pst DC3000.
Supplementary Figure S3. Callose and Camalexin deposition in 3h1
(A,B) Callose quantification in WT and 3h1 seedlings before and after 24 h flg22 treatment.
(C,D) Camalexin quantification in WT and 3h1 plants is shown as ng/g of fresh weight.
Supplementary Figure S4. (A) qRT-PCR validation of indicated defense related genes which showed altered expression pattern between WT and 3h1 before and after infection when pre-treated with flg22.
(B,C) qRT-PCR expression analysis of histone acetylation regulators HAC2 and HDA18 between WT and 3h1 before and after infection when pre-treated with flg22.
TUB6 was used for normalization.
Supplementary Figure S5.
(A-C) Distribution of chloroplast DNA methylation in all three sequence contexts in WT, WT + flg22, 3h1 and 3h1 + flg22.
(D-F) Percentage of non-conversion in all three sequence contexts
Supplementary Figure S6.
(A-C) Principal component analysis (PCA) plots of WGBS data of WT and 3h1 plants before and after flg22 treatment in all three (A) mCG (B) mCHG and (C) mCHH methylation contexts.
(D-F) Heatmap of Pearson correlation coefficient between all the samples
(G-I) Boxplots of mean DNA methylation in WT, WT + flg22, 3h1 and 3h1 + flg22 for individual biological replicates.
(J-L) Boxplots of average DNA methylation in WT, WT + flg22, 3h1 and 3h1 + flg22 in all three sequence contexts.
Supplementary Figure S7.
(A-C) Boxplots of mean DNA methylation in WT, WT + flg22, 3h1 and 3h1 + flg22 for individual biological replicates. Data represents methylation percentage for 2 Kb upstream prompter (A), gene body (B) and TEs (C) regions in all three contexts CG, CHG and CHH. p values represent Wilcoxon paired test.
Supplementary Figure S8.
DNA methylation dynamics of 154 genes of Cluster 1 (Figure 4) in 3h1 plants after flg22 treatment
(A-C) Heatmap of mean promoter methylation in all contexts (CG, CHG and CHH) of 154 genes in Cluster 1 in flg22-treated WT and 3h1 samples which showed differential expression after Pst DC3000 infection (Fig 4D).
(D-F) Box plot of mean promoter methylation in all contexts (CG, CHG and CHH) of Cluster 1 (154 genes) in flg22-treated WT and 3h1 samples which showed differential expression after Pst DC3000 infection (Cluster 1 of Fig 4D). p values represent Wilcoxon test.
Supplementary Figure S9. Western blot showing the H3K4me3 and H3K27me3 levels in WT and 3h1 plants after water (mock) or flg22 treatment challenged with Pst DC3000.
Supplementary Figure S10.
(A) DNase I accessibility PCR of HDA18 and WRKY62 in WT as compared to 3h1.
(B, C) DNase I accessibility PCR of HAC2 and WRKY29 in WT as compared to 3h1 before and after flg22 treatment.
Supplementary Table 1. Table showing the number of raw reads, trimmed reads, mapped reads, and conversion efficiency (by checking the % methylation of reads that map to the chloroplast genome) for each library. The non-conversion rates are indicated as percent methylation in CG, CHG and CHH contexts respectively
Gene
Name F R
FRK1 ACGGGCATAGTTCCACAAAG CGTCAAAAGAACGACGATGA PR1 CTCATACACTCTGGTGGG TTGGCACATCCGAGTC WRKY29 GCGTAAATACGGGCAGAAAC GGTTTGGGTTGGGAAGTTTT MYB51 ACAAATGGTCTGCTATAGCT CTTGTGTGTAACTGGATCAA RBOHD CCGAAGGTCCTTATCGACGG GTTCTCAATGTCGCTGTCGC RBOHF TGACACGCCAAGACGAAAGA GAGCAGAACGAGCATCACCT HDA18 AGAAGCCGAGTTGGGAATGG TTGTGGGAACGCTCCATGTT HAC2 AGGTGAACGGCGTACTGTTT GCTCGGTAATCCGGACAGAG WRKY38 CGCTCGAACGGTTTCTCCTT ATCCCACGAGTCTGGGTTGT WRKY62 GATCTACCACGACGGCTTCC ATCCCACGAGTCTGGGTTGT MAPKKK15 GGGTTGTGCTAAAACGGTGG ACCCCAAAGCCCAAACATCA PUB22 AGTTGCGGATCCAATGCAGA TGCTGATCTTGTCGCCTTGT EXO70H1 TCTGCGCTTATGCTGGGAAT GCTTCCTCAAATGCCGTGTG WRKY31 GCTCCTCAAGCATGGCTACA AGGGTTGTTACCGTTGGGTG SIF2 GGTCTTGTTGGGTACTGCGA CCGGTTACGTGTTCCTGACA MYB15 TTTCAAAACTTGGGCTCCGC CGCCGGTTCTAGCCAATACA MVQ2 ACATCAACCCGACCCATTCC TGTCAGGGCTGAGATCGAGA TUB6 GTCATCTGCAGTTGCGTCTT GGTGAAGGAATGGACGAGAT
Supplementary Table 2. Primer list used for RT-qPCR analysis in this study
