SlMBD5 Promotes Seed Germination by Epigenetically Repressing the Gibberellin Catabolism Gene SlGA2ox4 in Tomato [ChIP-Seq]

Seed germination represents a critical developmental transition in spermatophytes. Gibberellins (GAs) are essential phytohormones regulating seed germination. GA2-oxidases (GA2ox) play a key role in modulating GA levels by converting active GAs to inactive forms, critically influencing seed development and germination. How seed germination is epigenetically regulated by DNA methylation is poorly understood. Here, we identified the tomato methyl-CpG-binding domain (MBD) protein SlMBD5 as a regulator of seed germination. The slmbd5 mutant exhibits delayed germination and reduced GA4/GA7 levels, which can be rescued by exogenous GA4+7 application. Transcriptomic and biochemical analyses revealed that SlMBD5 represses the GA catabolism gene SlGA2ox4 by directly binding to its hypermethylated promoter. Furthermore, we show that SlMBD5 interacts with the histone methylation reader SlEBS, forming a functional complex that promotes the transcriptional repression of SlGA2ox4. Consistent with the model that SIMBD5 promotes seed germination through its repression of SlGA2ox4, the slmbd5/slga2ox4 double mutant shows partially restored germination. This study thus reveals an SlMBD5-SlEBS module that epigenetically regulates GA homeostasis to modulate seed germination in tomato. Overall design: ChIP-seq

种子萌发是种子植物（Spermatophytes）关键的发育转变过程。赤霉素（Gibberellins, GAs）是调控种子萌发的必需植物激素。GA2-氧化酶（GA2-oxidases, GA2ox）可通过将活性赤霉素转化为非活性形式来调控赤霉素水平，对种子发育与萌发具有关键影响。目前，DNA甲基化如何通过表观遗传机制调控种子萌发的相关机制仍不甚明晰。本研究鉴定到番茄的甲基-CpG结合域（methyl-CpG-binding domain, MBD）蛋白SlMBD5作为种子萌发的调控因子。slmbd5突变体表现为种子萌发延迟且GA4/GA7水平降低，该表型可通过外源施加GA4+7得以挽救。转录组学与生化分析显示，SlMBD5可直接结合赤霉素代谢基因SlGA2ox4的高甲基化启动子，从而抑制其转录。进一步研究表明，SlMBD5可与组蛋白甲基化阅读器SlEBS发生互作，形成功能复合体以增强对SlGA2ox4的转录抑制。与"SlMBD5通过抑制SlGA2ox4促进种子萌发"的模型一致，slmbd5/slga2ox4双突变体的种子萌发能力得到部分恢复。综上，本研究揭示了SlMBD5-SlEBS模块可通过表观遗传调控赤霉素稳态，进而调控番茄的种子萌发。整体实验设计：染色质免疫共沉淀测序（ChIP-seq）