Grassland Agri-Husbandry Research Center, College of Grassland Science, Qingdao Agricultural University, Qingdao, China; Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing, 100193, China.
Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing, 100193, China.
J Plant Physiol. 2024 Apr;295:154207. doi: 10.1016/j.jplph.2024.154207. Epub 2024 Feb 28.
Alfalfa (Medicago sativa) is one of the most widely cultivated forage crops in the world. However, alfalfa yield and quality are adversely affected by salinity stress. Nodulin 26-like intrinsic proteins (NIPs) play essential roles in water and small molecules transport and response to salt stress. Here, we isolated a salt stress responsive MsNIP2 gene and demonstrated its functions by overexpression in alfalfa. The open reading frame of MsNIP2 is 816 bp in length, and it encodes 272 amino acids. It has six transmembrane domains and two NPA motifs. MsNIP2 showed high identity to other known NIP proteins, and its tertiary model was similar to the crystal structure of OsNIP2-1 (7cjs) tetramer. Subcellular localization analysis showed that MsNIP2 protein fused with green fluorescent protein (GFP) was localized to the plasma membrane. Transgenic alfalfa lines overexpressing MsNIP2 showed significantly higher height and branch number compared with the non-transgenic control. The POD and CAT activity of the transgenic alfalfa lines was significantly increased and their MDA content was notably reduced compared with the control group under the treatment of NaCl. The transgenic lines showed higher capability in scavenging oxygen radicals with lighter NBT staining than the control under salt stress. The transgenic lines showed relative lower water loss rate and electrolyte leakage, but relatively higher Na content than the control line under salt stress. The relative expression levels of abiotic-stress-related genes (MsHSP23, MsCOR47, MsATPase, and MsRD2) in three transgenic lines were compared with the control, among them, only the expression of MsCOR47 was up-regulated. Consequently, this study offers a novel perspective for exploring the function of MsNIP2 in improving salt tolerance of alfalfa.
紫花苜蓿（Medicago sativa）是世界上栽培最广泛的饲料作物之一。然而，紫花苜蓿的产量和品质受到盐胁迫的不利影响。类硝酸还原酶 26 样内在蛋白（NIPs）在水和小分子的运输以及对盐胁迫的响应中发挥着重要作用。本研究中，我们分离了一个盐胁迫响应的 MsNIP2 基因，并通过在紫花苜蓿中的过表达证明了其功能。MsNIP2 的开放阅读框长 816bp，编码 272 个氨基酸。它有六个跨膜结构域和两个 NPA 基序。MsNIP2 与其他已知的 NIP 蛋白具有高度的同源性，其三级结构与 OsNIP2-1（7cjs）四聚体的晶体结构相似。亚细胞定位分析表明，与绿色荧光蛋白（GFP）融合的 MsNIP2 蛋白定位于质膜。与非转基因对照相比，过表达 MsNIP2 的转基因紫花苜蓿株系的株高和分枝数显著增加。在 NaCl 处理下，过表达 MsNIP2 的转基因紫花苜蓿株系的 POD 和 CAT 活性显著升高，MDA 含量显著降低，而对照组则相反。在盐胁迫下，与对照组相比，转基因株系的氧自由基清除能力更高，NBT 染色较轻。在盐胁迫下，转基因株系的相对失水量和电解质渗漏率较低，而 Na 含量相对较高。与对照相比，三个转基因株系中与非生物胁迫相关基因（MsHSP23、MsCOR47、MsATPase 和 MsRD2）的相对表达水平有所不同，其中只有 MsCOR47 的表达上调。因此，本研究为探索 MsNIP2 提高紫花苜蓿耐盐性的功能提供了新的视角。
