SENSITIVE TO PROTON RHIZOTOXICITY 1 (STOP1) is a core transcription factor that regulates the expression of aluminum (Al)-resistance genes to manage Al toxicity in plants. However, the genome-wide roles of SlSTOP1 in the Al stress response of tomato (Solanum lycopersicum) remain largely unknown. Here, we report that SlSTOP1 is crucial for Al tolerance in tomato, as loss-of-function mutants of SlSTOP1 displayed hypersensitivity to Al stress. Aluminum stress had no effect on SlSTOP1 mRNA expression, but promoted accumulation of SlSTOP1 protein in the nucleus. Through integrated DNA affinity purification sequencing and RNA sequencing analysis, we identified 39 SlSTOP1-targeted Al-responsive genes, some of which are homologous to known Al resistance genes in other plant species, suggesting that these SlSTOP1-targeted genes play essential roles in Al resistance in tomato. Furthermore, using peak enrichment analysis of SlSTOP1-targeted sequences, we identified a cis-acting element bound by SlSTOP1 and validated this finding via dual-luciferase reporter and electrophoretic mobility shift assay (EMSA). Additionally, we demonstrated SlHAK5 is one of direct targets of SlSTOP1 and functionally characterized it in terms of Al stress tolerance. Compared with wild-type plants, Slhak5 mutants developed by CRISPR/Cas9 technology presented increased sensitivity to Al stress, which was associated with reduced citrate secretion from the roots. Together, our findings demonstrate that SlSTOP1 directly interacts with cis-acting elements located in the promoters of target genes involved in diverse pathways contributing to Al resistance in tomato.

中文翻译：

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SlSTOP1 调节的 SlHAK5 表达通过促进根部分泌柠檬酸盐赋予番茄对 Al 的耐受性


对质子根茎毒性敏感 1 （STOP1） 是一种核心转录因子，可调节铝 （Al） 抗性基因的表达以管理植物中的 Al 毒性。然而，SlSTOP1 在番茄 （Solanum lycopersicum） 的 Al 应激反应中的全基因组作用在很大程度上仍然未知。在这里，我们报道了 SlSTOP1 对番茄的 Al 耐受性至关重要，因为 SlSTOP1 的功能丧失突变体对 Al 应激表现出超敏反应。铝胁迫对 SlSTOP1 mRNA 表达没有影响，但促进了 SlSTOP1 蛋白在细胞核中的积累。通过整合 DNA 亲和纯化测序和 RNA 测序分析，我们鉴定了 39 个 SlSTOP1 靶向的 Al 反应基因，其中一些与其他植物物种中已知的 Al 抗性基因同源，表明这些 SlSTOP1 靶向基因在番茄 Al 抗性中起着重要作用。此外，使用 SlSTOP1 靶向序列的峰富集分析，我们鉴定了与 SlSTOP1 结合的顺式作用元件，并通过双荧光素酶报告基因和电泳迁移率变化测定 （EMSA） 验证了这一发现。此外，我们证明了 SlHAK5 是 SlSTOP1 的直接靶标之一，并根据 Al 应力耐受性对其进行功能表征。与野生型植物相比，通过 CRISPR/Cas9 技术开发的 Slhak5 突变体对 Al 胁迫的敏感性增加，这与根部柠檬酸盐分泌减少有关。总之，我们的研究结果表明，SlSTOP1 直接与位于靶基因启动子中的顺式作用元件相互作用，这些靶基因参与导致番茄 Al 耐药性的不同途径。