Department of Plant Biology, Uppsala BioCenter, Swedish University of Agricultural Sciences and Linnean Centre for Plant Biology, Uppsala 75007, Sweden.
Nucleic Acids Res. 2021 May 7;49(8):4371-4385. doi: 10.1093/nar/gkab191.
Higher-order chromatin structure undergoes striking changes in response to various developmental and environmental signals, causing distinct cell types to adopt specific chromatin organization. High throughput chromatin conformation capture (Hi-C) allows studying higher-order chromatin structure; however, this technique requires substantial amounts of starting material, which has limited the establishment of cell type-specific higher-order chromatin structure in plants. To overcome this limitation, we established a protocol that is applicable to a limited amount of nuclei by combining the INTACT (isolation of nuclei tagged in specific cell types) method and Hi-C (INT-Hi-C). Using this INT-Hi-C protocol, we generated Hi-C data from INTACT purified endosperm and leaf nuclei. Our INT-Hi-C data from leaf accurately reiterated chromatin interaction patterns derived from conventional leaf Hi-C data. We found that the higher-order chromatin organization of mixed leaf tissues and endosperm differs and that DNA methylation and repressive histone marks positively correlate with the chromatin compaction level. We furthermore found that self-looped interacting genes have increased expression in leaves and endosperm and that interacting intergenic regions negatively impact on gene expression in the endosperm. Last, we identified several imprinted genes involved in long-range and trans interactions exclusively in endosperm. Our study provides evidence that the endosperm adopts a distinct higher-order chromatin structure that differs from other cell types in plants and that chromatin interactions influence transcriptional activity.
高等染色质结构会对各种发育和环境信号发生显著变化，使不同的细胞类型采用特定的染色质组织。高通量染色质构象捕获（Hi-C）允许研究高等染色质结构；然而，这项技术需要大量的起始材料，这限制了在植物中建立特定细胞类型的高等染色质结构。为了克服这个限制，我们通过结合 INTACT（特定细胞类型中标记的核的分离）方法和 Hi-C（INT-Hi-C）建立了一种适用于有限量核的方案。使用这种 INT-Hi-C 方案，我们从 INTACT 纯化的胚乳和叶片核中生成了 Hi-C 数据。我们从叶片获得的 INT-Hi-C 数据准确地重复了来自常规叶片 Hi-C 数据的染色质相互作用模式。我们发现，混合叶片组织和胚乳的高等染色质组织不同，并且 DNA 甲基化和抑制性组蛋白标记与染色质紧缩水平呈正相关。我们还发现，自我环化的相互作用基因在叶片和胚乳中表达增加，并且相互作用的基因间区域对胚乳中的基因表达有负面影响。最后，我们在胚乳中鉴定了几个涉及长距离和转座相互作用的印迹基因。我们的研究提供了证据，证明胚乳采用了与植物中其他细胞类型不同的独特的高等染色质结构，并且染色质相互作用影响转录活性。
