Key message

OsSIRP4 is an E3 ligase that acts as a negative regulator in the plant response to salt stress via the 26S proteasomal system regulation of substrate proteins, OsPEX11-1, which it provides important information for adaptation and regulation in rice.

Abstract

Plants are sessile organisms that can be exposed to environmental stress. Plants alter their cellular processes to survive under potentially unfavorable conditions. Protein ubiquitination is an important post-translational modification that has a crucial role in various cellular signaling processes in abiotic stress response. In this study, we characterized Oryza sativa salt-induced RING finger protein 4, OsSIRP4, a membrane and cytosol-localized RING E3 ligase in rice. OsSIRP4 transcripts were highly induced under salt stress in rice. We found that OsSIRP4 possesses E3 ligase activity; however, no E3 ligase activity was observed with a single amino acid substitution (OsSIRP4^C269A). The results of the yeast two hybrid system, in vitro pull-down assay, BiFC analysis, in vitro ubiquitination assay, and in vitro degradation assay indicate that OsSIRP4 regulates degradation of a substrate protein, OsPEX11-1 (Oryza sativa peroxisomal biogenesis factor 11-1) via the 26S proteasomal system. Phenotypic analysis of OsSIRP4-overexpressing plants demonstrated hypersensitivity to salt response compared to that of the wild type and mutated OsSIRP4^C269A plants. In addition, OsSIRP4-overexpressing plants exhibited significant low enzyme activities of superoxide dismutase, catalase, and peroxidase, and accumulation of proline and soluble sugar, but a high level of H₂O₂. Furthermore, qRT data on transgenic plants suggest that OsSIRP4 acted as a negative regulator of salt response by diminishing the expression of genes related to Na⁺/K⁺ homeostasis (AtSOS1, AtAKT1, AtNHX1, and AtHKT1;1) in transgenic plants under salt stress. These results suggest that OsSIRP4 plays a negative regulatory role in response to salt stress by modulating the target protein levels.

中文翻译：

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E3 连接酶，即稻盐诱导的环指蛋白 4 (OsSIRP4)，通过降解 OsPEX11-1 蛋白负调节盐胁迫反应

关键信息

OsSIRP4是一种E3连接酶，通过26S蛋白酶体系统调控底物蛋白OsPEX11-1作为植物对盐胁迫响应的负调控因子，为水稻的适应和调控提供重要信息。

抽象的

植物是可以暴露于环境压力的固着生物。植物改变它们的细胞过程以在潜在的不利条件下生存。蛋白质泛素化是一种重要的翻译后修饰，在非生物应激反应的各种细胞信号传导过程中起着至关重要的作用。在这项研究中，我们对水稻盐诱导的 RING 指蛋白 4、OsSIRP4 进行了表征，这是一种水稻膜和胞质溶胶定位的 RING E3 连接酶。OsSIRP4 转录本在水稻盐胁迫下被高度诱导。我们发现OsSIRP4具有E3连接酶活性；然而，没有观察到单个氨基酸取代的 E3 连接酶活性（OsSIRP4 ^C269A）。酵母二杂交系统、体外下拉试验、BiFC 分析、体外泛素化试验和体外降解试验的结果表明，OsSIRP4 调节底物蛋白 OsPEX11-1 ( Oryza sativa peroxisomal biogenesis factor 11- 1) 通过 26S 蛋白酶体系统。与野生型和突变的 OsSIRP4 ^C269A植物相比，过表达 OsSIRP4 的植物的表型分析表明对盐反应的超敏反应。此外，过表达OsSIRP4的植物的超氧化物歧化酶、过氧化氢酶和过氧化物酶的酶活性显着降低，脯氨酸和可溶性糖积累，但H ₂ O _{2水平较高。}. 此外，转基因植物的 qRT 数据表明，OsSIRP4 通过减少盐胁迫下转基因植物中与 Na ⁺ /K ⁺稳态相关的基因（AtSOS1、AtAKT1、AtNHX1和AtHKT1;1）的表达，作为盐反应的负调节因子。 . 这些结果表明，OsSIRP4 通过调节靶蛋白水平在盐胁迫响应中发挥负调节作用。