Fig. 3: Determination of DNA-binding sites and target genes of CaKAN3/CaHSF8 by ChIP-seq.

a Genome-wide distribution of DNA-binding peaks of CaKAN3/CaHSF8. b The reads per million (RPM) of CaHSF8, CaKAN3 and input are shown as a heatmap, and 199 cotargeted genes by CaHSF8/CaKAN3 are shown as Venn diagrams. c The genes cotargeted by CaHSF8/CaKAN3 were enriched in different KEGG signaling pathways. d Integrative Genomics Viewer (IGV) images of ChIP-seq data and the locations of HSE and AACAA motifs within the promoters of the CaR1B23, CaR1B11, CaR1A, CaR1B12, CaR1A6 and CaR1B-16 genes that are cotargeted by CaHSF8/CaKAN3. e Both CaKAN3 and CaHSF8 exhibited higher levels of enrichment on the promoters of the 6 tested NLR genes. GV3101 cells containing 35 S:CaKAN3-HA and 35 S:CaHSF8-HA were infiltrated into pepper leaves, which were harvested at 48 hpi for ChIP‒qPCR analysis using specific primer pairs. IP using IgG beads was used as the control. The enrichment levels of the tested genes were compared with those in the control, and the relative enrichment of IP using anti-HA was set to a value of 1 after normalization by input. Data are shown as the means ± standard errors of three replicates. Asterisks above the bars indicate significant differences between means (P < 0.01), as calculated with Fisher’s protected t test. CaWRKY40 and CaWRKY58 were used as negative controls. The ratio of IP:anti-HA to IP:IgG is indicated on the error line of IP:IgG. All replicates were from different plants. Source data are provided as a Source Data file. f CaKAN3-GST and CaHSF8-GST bound the promoters of the 6 NLR genes in an AACAA-element- and HSE-dependent manner, as shown by EMSA. The experiment was carried out twice with similar results.