番茄乙烯响应因子SlERF.F12抑制果实成熟的分子机制

The gaseous plant hormone ethylene plays a key role in fruit ripening, but the role and mode of action of Ethylene Response Factors (ERFs) in fruit ripening remain poorly understood. Our earlier research found that SlERF.F12, a member of ERF.F subfamily that act as transcriptional repressors, displayed a reduced expression during ripening process. Preliminary studies showed that mutation of SlERF.F12 by the CRISPR/Cas9 gene editing system resulted in an accelerated fruit ripening and overexpression of SlERF.F12 delayed fruit ripening, suggesting a involvement of this gene in repression of fruit ripening. Nevertheless, the molecular mechanism of SlERF.F12 in controlling fruit ripening is yet to be elucidated. This project aims to investigate the role of SlERF.F12 in fruit ripening through integrated phenotypical, physiological, biochemical, and molecular analysis of the SlERF.F12 overexpression lines and erf.f12 mutants. To reveal the molecular mechanisms of SlERF.F12 in controlling fruit ripening and its downstream regulatory networks, the target genes of SlERF.F12 will be identified by integrated RNA-seq, ChIP-seq, EMSA and protoplast transient expression system. The outcomes of this project are of great significance for demonstration of the function and molecular mechanisms of downstream components of ethylene signaling in controlling fruit ripening.

植物激素乙烯在果实成熟过程中发挥关键作用。目前对乙烯信号途径最下游的乙烯响应因子（ERFs）在果实成熟过程中的功能及调控机制还不完全清楚。我们前期研究发现转录调控中起抑制作用的ERF.F亚家族基因SlERF.F12在果实成熟过程中特异性下调表达，且超表达SlERF.F12导致果实成熟显著推迟，而通过CRISPR/Cas9基因编辑获得的突变体中果实成熟明显提前。这些结果表明SlERF.F12在果实成熟过程中发挥抑制作用，但调控机制尚不清楚。因此，本项目拟通过对SlERF.F12超表达和突变体株系的表型鉴定明确SlERF.F12在果实成熟过程中的功能。并利用RNA-seq、ChIP-seq、EMSA以及转录激活等技术和方法鉴定SlERF.F12调控的靶基因，揭示SlERF.F12基因调控果实成熟的分子机制及其下游调控网络。研究结果对揭示果实成熟调控网络具有重要意义。

果实的成熟是一个高度程序化调控的复杂过程，受到外源环境和内源因子等多种因素的调控。对于番茄等呼吸跃变型果实，目前已知植物激素乙烯、关键转录因子以及表观遗传修饰在成熟过程中发挥重要作用，但对于它们之间的关联和协同机制还有待揭示。本项目从番茄ERF家族中筛选到一个受乙烯抑制且在果实成熟过程中特异性下调表达的SlERF.F12基因。SlERF.F12编码蛋白含有两个典型的转录抑制EAR基序，转录激活实验表明SlERF.F12依赖其C-末端EAR基序发挥转录抑制作用。对SlERF.F12过表达、RNAi以及基因编辑突变体的表型分析发现，SlERF.F12在抑制果实成熟启动、延长果实采后储存时间方面发挥重要功能。结合转录组、转录激活、EMSA、ChIP-qPCR等实验发现SlERF.F12直接抑制乙烯合成、细胞壁降解等成熟相关基因的转录。进一步研究显示，SlERF.F12依赖其C-末端EAR基序招募转录抑制辅因子TPL2和组蛋白去乙酰化酶HDA1/3组成转录抑制复合体，通过抑制乙烯合成、果实软化等基因启动子区域H3K9和H3K27位点的乙酰化修饰水平进而抑制成熟相关基因的表达。研究结果揭示了受乙烯抑制的转录因子SlERF.F12通过表观修饰调控果实成熟的分子机制，为果实贮藏保鲜技术的研发提供参考。