Plant Genomics Laboratory, Department of Bio-Resources Sciences, Graduate School, Kangwon National University, Chuncheon, 200-713, South Korea.
Plant Mol Biol. 2021 Feb;105(3):231-245. doi: 10.1007/s11103-020-01084-x. Epub 2020 Oct 20.
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. 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). 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 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 HO. 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.
OsSIRP4 是一种 E3 连接酶，通过 26S 蛋白酶体系统调节底物蛋白 OsPEX11-1 的表达，作为植物响应盐胁迫的负调控因子，为水稻适应和调控提供重要信息。植物是不能移动的生物，它们可能会暴露在环境压力下。植物会改变它们的细胞过程，以在潜在不利的条件下生存。蛋白质泛素化是一种重要的翻译后修饰，在非生物胁迫响应中的各种细胞信号过程中起着至关重要的作用。在这项研究中，我们对水稻中膜和细胞质定位的 RING E3 连接酶 OsSIRP4 进行了盐诱导的水稻盐诱导 RING 指蛋白 4（OsSIRP4）的特征描述。在水稻中，OsSIRP4 转录物在盐胁迫下高度诱导。我们发现 OsSIRP4 具有 E3 连接酶活性；然而，单个氨基酸替换（OsSIRP4）时没有观察到 E3 连接酶活性。酵母双杂交系统、体外下拉测定、BiFC 分析、体外泛素化测定和体外降解测定的结果表明，OsSIRP4 通过 26S 蛋白酶体系统调节底物蛋白 OsPEX11-1（Oryza sativa 过氧化物酶体生物发生因子 11-1）的降解。与野生型和突变 OsSIRP4 植物相比，OsSIRP4 过表达植物的表型分析显示对盐反应的敏感性增加。此外，OsSIRP4 过表达植物的超氧化物歧化酶、过氧化氢酶和过氧化物酶的活性显著降低，脯氨酸和可溶性糖的积累增加，但 HO 水平升高。此外，转基因植物的 qRT 数据表明，OsSIRP4 通过减少与 Na/K 稳态相关的基因（AtSOS1、AtAKT1、AtNHX1 和 AtHKT1；1）的表达，在盐胁迫下作为盐响应的负调控因子发挥作用。这些结果表明，OsSIRP4 通过调节靶蛋白水平在响应盐胁迫中发挥负调节作用。
