Nuclear lamina component KAKU4 regulates chromatin states and transcriptional regulation in the Arabidopsis genome

Abstract

Background: The nuclear lamina links the nuclear membrane to chromosomes and plays a crucial role in regulating chromatin states and gene expression. However, current knowledge of nuclear lamina in plants is limited compared to animals and humans.

Results: This study mainly focused on elucidating the mechanism through which the putative nuclear lamina component protein KAKU4 regulates chromatin states and gene expression in Arabidopsis leaves. Thus, we constructed a network using the association proteins of lamin-like proteins, revealing that KAKU4 is strongly associated with chromatin or epigenetic modifiers. Then, we conducted ChIP-seq technology to generate global epigenomic profiles of H3K4me3, H3K27me3, and H3K9me2 in Arabidopsis leaves for mutant (kaku4-2) and wild-type (WT) plants alongside RNA-seq method to generate gene expression profiles. The comprehensive chromatin state-based analyses indicate that the knockdown of KAKU4 has the strongest effect on H3K27me3, followed by H3K9me2, and the least impact on H3K4me3, leading to significant changes in chromatin states in the Arabidopsis genome. We discovered that the knockdown of the KAKU4 gene caused a transition between two types of repressive epigenetics marks, H3K9me2 and H3K27me3, in some specific PLAD regions. The combination analyses of epigenomic and transcriptomic data between the kaku4-2 mutant and WT suggested that KAKU4 may regulate key biological processes, such as programmed cell death and hormone signaling pathways, by affecting H3K27me3 modification in Arabidopsis leaves.

Conclusions: In summary, our results indicated that KAKU4 is directly and/or indirectly associated with chromatin/epigenetic modifiers and demonstrated the essential roles of KAKU4 in regulating chromatin states, transcriptional regulation, and diverse biological processes in Arabidopsis.

Keywords: Arabidopsis; Chromatin state; H3K27me3; H3K9me2; Hormone pathway; KAKU4.

Fig. 1

Protein-protein association network analysis of lamin-like proteins. A Protein-protein association network of lamin-like proteins. The orange nodes represent the lamin-like proteins; the yellow nodes represent the proteins related to chromatin or epigenetic modification; the gray nodes represent the other proteins. The line color represents the method of demonstrating protein-protein interactions: The dark blue line represents the experimental verification of protein-protein interaction, the light blue line represents MS verification of a single protein, the green line represents the identification of large-scale yeast two-hybrid, and the yellow line represents the identification of large-scale co-fractionation mass spectrometry (CF-MS). The thickness of the line represents the number of evidence. B GO enrichment analysis of lamin-like proteins and their possible associated proteins. The bar color represents the GO category: red indicates biological processes (BP), green indicates molecular functions (MF), and blue indicates cellular components (CC). C Chromatin or epigenetic modification proteins among associated proteins of lamin-like proteins

Fig. 2

ChIP-seq analysis for histone modification in the kaku4-2 mutant and WT. A The H3K4me3/H3K27me3/H3K9me2 distribution within different regions (promoter, 5′UTR, coding exon, intron, 3′UTR, and intergenic) of the Arabidopsis genome in kaku4-2 mutant and WT. B The heatmaps display the H3K4me3/H3K27me3/H3K9me2 signal around TSSs in kaku4-2 mutant and WT plants. For each gene, the H3K4me3/H3K27me3/H3K9me2 signals are displayed along −1 to 1 kb regions around the TSSs. C1–C8 are the eight parts after clustering. C Distribution of H3K4me3/H3K27me3/H3K9me2 along Arabidopsis genes in the C1 region. A meta-gene profile was generated using the normalized sequencing density of H3K4me3/H3K27me3/H3K9me2 in kaku4-2 mutant and WT. The gene body was converted into a percentage to standardize genes of different lengths. The 1 kb upstream and downstream regions of the gene are included. D Distribution of H3K4me3/H3K27me3/H3K9me2 along Arabidopsis genes in the C6 region. A meta-gene profile was generated using the normalized sequencing density of H3K4me3/H3K27me3/H3K9me2 in kaku4-2 mutant and WT

Fig. 3

Chromatin state analysis for differential histone modification deposition between the kaku4-2 mutant and WT. A Chromatin state analysis for the regions with higher deposition of H3K4me3, H3K27me3, and H3K9me2 in kaku4-2 mutant and WT plants. The upper plot X-axis represents the 36 chromatin states, and the Y-axis represents the proportion of chromatin states enriched in the regions with higher H3K4me3, H3K27me3, and H3K9me2 deposition in the kaku4-2 mutant and WT plants; red bars represent higher deposition of H3K4me3, H3K27me3, and H3K9me2 in kaku4-2 mutant; the blue bars represent higher deposition of H3K4me3, H3K27me3, and H3K9me2 in the WT. ***P < 0.001, **P < 0.01, *P < 0.05. The lower panels represent a heatmap of the enriched epigenetic marks in each chromatin state. Color-filled indicates preferential epigenetic marks, while no color fill represents the absence or low signal of the indicated epigenetic mark. B Enrichment profiles of H3K4me3/H3K27me3/H3K9me2 deposition in PLADs. A profile was generated using the normalized sequencing density of H3K4me3/H3K27me3/H3K9me2 in kaku4-2 mutant and WT. PLADs of different lengths are standardized to 3 kb. The 2 kb upstream and downstream regions of the PLAD are included

Fig. 4

Combination analyses of epigenomic and transcriptomic data of the kaku4-2 mutant and WT. A Heatmap showing relative expression levels of differentially expressed genes in different replicates of the kaku4-2 mutant and WT. B GO enrichment analyses for genes upregulated in kaku4-2 mutant by agriGOv2 and REVIGO. The scatter plot shows the cluster representatives in two-dimensional space derived by applying multidimensional scaling to a matrix of the significant GO terms with semantic similarities. The color and size of the bubble indicate the log₁₀FDR (a legend in the bottom right-hand corner). The colors from red to green represent the significance level of the GO terms, from high to low. Fisher's exact test was conducted, and the P-value was adjusted using the Benjamini-Yekutieli method, FDR ≤ 0.05. C GO enrichment analyses for genes downregulated in kaku4-2 mutant by agriGOv2 and REVIGO. D GO enrichment analyses for genes upregulated in the kaku4-2 mutant and with higher deposition of H3K27me3 in the WT by agriGO and REVIGO