Zhejiang Key Laboratory of Crop Germplasm, College of Agriculture & Biotechnology, Zhejiang University, Hangzhou, 310058, China.
Zhejiang Key Laboratory of Crop Germplasm, College of Agriculture & Biotechnology, Zhejiang University, Hangzhou, 310058, China; Shandong (Linyi) Institute of Modern Agriculture, Zhejiang University, Linyi, 276000, China.
Plant Physiol Biochem. 2022 Dec 15;193:110-123. doi: 10.1016/j.plaphy.2022.10.033. Epub 2022 Nov 3.
The most hazardous abiotic stress, salinity, restricted the world crop production, and grain chalkiness affected the grain quality to limit consumers' acceptance. The basic helix-loop-helix (bHLH) proteins modulate massive biological processes in plants. Here the CRISPR/Cas9 gene editing mutants were obtained to detect the function of OsbHLH044. The loss-of-function of OsbHLH044 mutants showed numerous altered plant phenotypes. Notably, the osbhlh044 mutants resulted in prominently reduced morphological and physiological parameters under salt stress. Lower antioxidant activities and higher lipid peroxidation and hydrogen peroxide (HO) accumulation in the osbhlh044 mutants caused salinity sensitivity due to elevated reactive oxygen species (ROS). Under salt stress, both shoots and roots of the osbhlh044 mutants acquired higher Na. Moreover, the expression of ion homeostasis-related genes (OsHKTs, OsHAK, OsSOSs, and OsNHX) and ABA-responsive gene (OsLEA3) was significantly altered in the osbhlh044 mutants after salt stress. The expression levels of genes coding for starch (OsAGPL1, OsSSIIa, OsWx, and OsFLO2) and seed storage proteins (GluA1 and Globulin 1) were significantly decreased, indicating that they synthesize less store starch and proteins, resulting in grain chalkiness in the osbhlh044 mutants. Yeast one Hybrid (Y1H) showed that OsbHLH044 could activate salt- (OsHKT1;3, OsHAK7, OsSOS1, OsSOS2, OsNHX2, and OsLEA3 but not OsHKT2;1), and starch-related genes (OsSSIIa, OsWx, and OsFLO2) by binding to the G-boxes of their promoters. Therefore, the OsbHLH044 gene editing mutants revealed multiple functions, specifically a positive regulator of salt stress and grain quality, which might bring new insights into the breeding of rice varieties.
最具危害性的非生物胁迫因素是盐度，它限制了世界作物的生产，而籽粒垩白度则影响了谷物的品质，限制了消费者的接受程度。碱性螺旋-环-螺旋（bHLH）蛋白调节着植物中的大量生物学过程。在这里，我们通过 CRISPR/Cas9 基因编辑获得突变体，以检测 OsbHLH044 的功能。OsbHLH044 突变体的功能丧失表现出许多改变的植物表型。值得注意的是，OsbHLH044 突变体在盐胁迫下导致形态和生理参数明显降低。OsbHLH044 突变体中较低的抗氧化活性和较高的脂质过氧化作用以及过氧化氢（HO）积累导致盐敏感性，这是由于活性氧（ROS）的升高所致。在盐胁迫下，osbhlh044 突变体的根和茎都积累了更高的 Na。此外，在盐胁迫后，离子稳态相关基因（OsHKTs、OsHAK、OsSOSs 和 OsNHX）和 ABA 响应基因（OsLEA3）的表达在 osbhlh044 突变体中发生了显著改变。编码淀粉（OsAGPL1、OsSSIIa、OsWx 和 OsFLO2）和种子贮藏蛋白（GluA1 和球蛋白 1）的基因的表达水平显著降低，表明它们合成的贮藏淀粉和蛋白质较少，导致 osbhlh044 突变体中的籽粒垩白度增加。酵母单杂交（Y1H）表明，OsbHLH044 可以通过结合其启动子中的 G 框来激活盐（OsHKT1;3、OsHAK7、OsSOS1、OsSOS2、OsNHX2 和 OsLEA3，但不是 OsHKT2;1）和淀粉相关基因（OsSSIIa、OsWx 和 OsFLO2）。因此，OsbHLH044 基因编辑突变体揭示了多种功能，特别是作为盐胁迫和籽粒品质的正调节剂，这可能为水稻品种的培育带来新的启示。
