拟南芥组蛋白H3K4去甲基化酶JMJ17调控干旱胁迫应答的分子机制

Emerging evidences indicate that the fluctuation of H3K4 methylation status directly links to gene expressional regulatory dynamics under the drought stress, thereby facilitating the plant short-term stress memory as well as the long-term stress adaptation. However, thus far, the specific histone demethylases which directly affect H3K4 levels in plant under the drought stress remains unknown. In the previous pilot experiment, we identified that evolutionarily conserved Jumonji domain-containing protein 17 (JMJ17) belonging to KDM5/JARID1 family functioned in the drought stress tolerance. jmj17 loss-of-function mutants displayed the higher drought stress tolerance compared to that of wild type plants (WT). This result indicates that JMJ17 acts as the negative regulator under the drought stress response. Further in vitro and in vivo histone demethylation assays revealed that JMJ17 specifically demethylated histone H3K4me1, H3K4me2 and H3K4me3. In the future work, we are going to investigate molecular regulatory mechanisms by which JMJ17 specifically regulates drought stress responsive gene expression. We would perform the RNA-sequencing analysis to identify the transcriptional downstream targets of JMJ17 under the drought stress. Further high throughput ChIP-sequencing analyses using H3K4me1, H3K4me2 and H3K4me3 specific antibodies in WT and jmj17 will provide us with the clues of how JMJ17 specifically regulates H3K4 methylation status under the drought stress. Moreover, we will perform ChIP-sequencing analysis using WT and pJMJ17:JMJ17-FLAG transgenic plants to identify direct downstream targets of JMJ17. Lastly, to investigate JMJ17 interacting partners, we will take advantages of LC-MS/MS technology and yeast two-hybrid screening methods, and will further examine the genetic interactions. Our study will shed new light on a novel molecular epigenetic regulatory mechanisms under the drought stress response.

在干旱胁迫条件下，关键胁迫应答基因位点的组蛋白H3K4甲基化水平直接影响其表达水平，从而决定植物对胁迫的适应程度。然而，特异性的组蛋白H3K4去甲基化酶尚未被发现。在前期工作中，我们发现，拟南芥jmj17突变体耐旱性增强，说明JMJ17负向调控拟南芥对干旱胁迫的应答。并且，JMJ17具有H3K4特异性去甲基化酶活性。我们拟利用染色质免疫共沉淀测序和RNA测序技术，在全基因组水平鉴定受JMJ17诱导的H3K4去甲基化作用调控的干旱胁迫应答基因，并找出JMJ17调控的重要的下游靶基因，进行遗传学互作验证 (genetic interaction)。同时，利用免疫共沉淀(Co-IP)结合LC-MS/MS技术和酵母双杂交系统，筛选JMJ17的互作蛋白，深入研究JMJ17在干旱胁迫应答过程中调控转录的分子机制。研究结果将为揭示植物干旱胁迫应答的表观遗传分子机制奠定基础。

气候变化正在加剧全球干旱，严重威胁着粮食安全。由于植物是着地固定生长，不能主动逃避环境危害，为了生存植物进化出复杂且精细的调控机制来减少干旱危害。因此解析植物的抗旱分子机制对于培育新的抗旱品种有重要的指导意义。组蛋白的修饰是生物表观遗传调控的一种重要方式，组蛋白的修饰改变了染色质的构型，进而影响基因转录。本研究发现拟南芥中的一个组蛋白H3第4位赖氨酸（H3K4）去甲基化酶，JMJ17，在干旱胁迫和脱落酸（ABA）应答途径中行使重要功能。本研究分析了组蛋白的去甲基化在干旱胁迫和ABA应答中的作用，阐明了H3K4去甲基化酶JMJ17调控干旱胁迫和ABA应答的分子机制。.本研究利用分子生物学、生物化学和生物信息分析等方法，系统地分析了组蛋白去甲基化酶JMJ17在干旱胁迫和ABA应答过程中的调控作用。主要的研究结果如下：与野生型相比，jmj17突变体表现出抗旱表型和对ABA敏感表型；利用体内、体外酶活实验，我们发现JMJ17有组蛋白去甲基化酶的活性，能够有效地降低H3K4me1/2/3水平；利用ChIP-seq和RNA-seq分析，发现在干旱胁迫后，与野生型相比，jmj17突变体中部分胁迫应答基因的转录水平与H3K4me3水平均显著上升，说明在干旱胁迫下JMJ17能影响H3K4me3水平同时还影响基因的表达；JMJ17与WRKY40互作，在ABA应答过程中调控共同靶基因；WRKY40对于JMJ17招募到ABI5染色质区域是必需的；ABA通过抑制WRKY40活性促进了HY5介导的ABI5激活；HY5和ABI5协同激活ABI5的表达。. 综上所述，本研究结果表明组蛋白去甲基化酶JMJ17，通过调控OST1，ABI5等胁迫应答基因的H3K4me3水平影响这些基因的表达，在干旱胁迫和ABA应答途径中行使负调控作用。