背景回顾：Enhancers located in introns are abundant and play a major role in the regulation of gene expression in mammalian species. 

提出问题：By contrast, the functions of intronic enhancers in plants have largely been unexplored and only a handful of plant intronic enhancers have been reported and characterized.

主要研究：We performed a genome-wide prediction of intronic enhancers in Arabidopsis thaliana using open chromatin signatures based on DNase I sequencing. 

结果1-鉴定：We identified 941 candidate intronic enhancers associated with 806 genes in seedling tissue and 1,271 intronic enhancers associated with 1,069 genes in floral tissue.

结果2-表达验证：We validated the function of 15 of 21 (71%) of the predicted intronic enhancers in transgenic assays using a reporter gene.

结果3-敲除验证：We also created deletion lines of three intronic enhancers associated with two different genes using CRISPR/Cas. Deletion of these enhancers, which span key transcription factor binding sites, did not abolish gene expression but caused varying levels of transcriptional repression of their cognate genes. Remarkably, the transcriptional repression of the deletion lines occurred at specific developmental stages and resulted in distinct phenotypic effects on plant morphology and development. 

结论：Clearly, these three intronic enhancers are important in fine-tuning tissue- and development-specific expression of their cognate genes.

 摘 要 

哺乳动物中，位于内含子区域的增强子十分丰富，并且在基因表达调控中发挥重要作用。相反，植物中内含区域的增强子功能还不明晰，目前仅报道并鉴定了少数几个植物内含子增强子的功能。本文中，作者利用基于DNase I测序的开放染色质信号检测，对拟南芥进行了全基因组范围上内含子增强子的预测。作者在实生苗组织中鉴定到了941个候选的内含子增强子，与806个基因相关；而在花组织中鉴定到了1271个候选的内含子增强子，与1069个基因相关。作者进一步对其中21个候选内含子增强子利用报告基因进行转基因验证，发现15个（约占71%）具有增强子功能。作者还通过CRISPR/Cas系统删除了与两个基因相关的三个内含子增强子，获得了突变体株系。这些增强子区域覆盖了关键转录因子的结合位点，而他们的删除并未测底导致关联基因不表达，但是确实导致了这些基因出现转录水平上不同程度的抑制。值得注意的是，增强子删除株系的转录抑制发生在特定的发育阶段，并且对植株的形态和发育起到明显的表型效应。显然，这三个内含子增强子在相关基因的组织和发育特异性表达过程中起到精细调控的作用。