DataSheet_1_Overexpression of stress granule protein TZF1 enhances salt stress tolerance by targeting ACA11 mRNA for degradation in Arabidopsis.pdf

Tandem CCCH zinc finger (TZF) proteins play diverse roles in plant growth and stress response. Although as many as 11 TZF proteins have been identified in Arabidopsis, little is known about the mechanism by which TZF proteins select and regulate the target mRNAs. Here, we report that Arabidopsis TZF1 is a bona-fide stress granule protein. Ectopic expression of TZF1 (TZF1 OE), but not an mRNA binding-defective mutant (TZF1H186Y OE), enhances salt stress tolerance in Arabidopsis. RNA-seq analyses of NaCl-treated plants revealed that the down-regulated genes in TZF1 OE plants are enriched for functions in salt and oxidative stress responses. Because many of these down-regulated mRNAs contain AU- and/or U-rich elements (AREs and/or UREs) in their 3’-UTRs, we hypothesized that TZF1—ARE/URE interaction might contribute to the observed gene expression changes. Results from RNA immunoprecipitation-quantitative PCR analysis, gel-shift, and mRNA half-life assays indicate that TZF1 binds and triggers degradation of the autoinhibited Ca2+-ATPase 11 (ACA11) mRNA, which encodes a tonoplast-localized calcium pump that extrudes calcium and dampens signal transduction pathways necessary for salt stress tolerance. Furthermore, this salt stress-tolerance phenotype was recapitulated in aca11 null mutants. Collectively, our findings demonstrate that TZF1 binds and initiates degradation of specific mRNAs to enhance salt stress tolerance.

串联CCCH锌指（Tandem CCCH zinc finger, TZF）蛋白在植物生长与胁迫响应中发挥多样功能。尽管拟南芥中已鉴定出多达11种TZF蛋白，但目前对TZF蛋白选择并调控靶标mRNA的分子机制仍知之甚少。本研究发现，拟南芥TZF1是一种真正的应激颗粒蛋白。异位表达TZF1（TZF1过表达，TZF1 OE）可增强拟南芥的盐胁迫耐受性，而mRNA结合缺陷型突变体（TZF1H186Y过表达，TZF1H186Y OE）则无此效果。对NaCl处理后的植株进行RNA测序（RNA-seq）分析发现，TZF1过表达植株中的下调基因富集于盐胁迫与氧化胁迫响应相关功能通路。由于这些下调mRNA的3'非翻译区（3'-UTR）大多含有AU富集元件（AREs）和/或U富集元件（UREs），我们推测TZF1与ARE/URE的相互作用可能促成了上述基因表达变化。RNA免疫沉淀定量PCR（RNA immunoprecipitation-quantitative PCR）、凝胶迁移实验以及mRNA半衰期测定的结果表明，TZF1可结合并启动自抑制型钙ATP酶11（autoinhibited Ca2+-ATPase 11, ACA11）mRNA的降解——该mRNA编码一种液泡膜定位的钙泵，可排出钙离子并抑制盐胁迫耐受所需的信号转导通路。此外，aca11缺失突变体重现了该盐胁迫耐受表型。综上，本研究结果证实，TZF1通过结合并启动特定mRNA的降解，从而增强植物的盐胁迫耐受性。