Genome-wide profiling and functional study of short N-terminal H2B variants in Arabidopsis.

INTRODUCTION: Nucleosomes harboring specific histone variants show distinct chromatin localization patterns and regulatory functions, thereby playing crucial roles in epigenetic regulation. Compared to the well-understood variants of H2A and H3, the study about H2B variants is emerging. Deciphering the roles and regulatory mechanisms of H2B variants in plants will provide more knowledges about epigenetic regulations in plant biology. OBJECTIVES: Using the model plant Arabidopsis thaliana as the research subject, we systematically analyzed histone H2B variants, four short N-terminal histone H2B variants (snH2Bs) were identified. The genomic distribution characteristics of these snH2Bs, their impact on plant growth, and the potential regulatory mechanisms were studied. METHODS: By integrating whole-genome chromatin immunoprecipitation sequencing (ChIP-seq) and fluorescence microscopy localization analysis, the distribution of snH2Bs across the genome was identified. Single, double, and triple knock-out mutants were constructed using CRISPR-Cas9 to further explore the functions of snH2Bs in the growth process of Arabidopsis thaliana, the possible mechanisms were also discussed. RESULTS: These snH2B variants are preferentially expressed in reproductive tissues and are detected in the nuclei of pollen grains. Further genome-wide profiling indicates that the snH2Bs distribute at active chromatin regions and are positively correlated with gene expression. By creating knock-out single, double, and triple mutants for these snH2Bs, we demonstrate that H2B.5 influences vegetative to reproductive transition. We also show that H2B.5 is required for proper accumulation of H3 lysine 9 acetylation and H2B mono-ubiquitination. CONCLUSION: Overall, our study not only provide insights into the functions and chromatin characteristics of plant snH2Bs, but also supplies examples that illustrate the interplay between histone variants and histone modification. These findings contribute to the understanding of the fundamental principles of epigenetic regulation in eukaryotes and also highlight potential targets for crop improvement.