Lv Guihua, Li Yunfeng, Wu Zhengxin, Zhang Yahui, Li Xiangnan, Wang Tingzheng, Ren Wenchuang, Liu Lei, Chen Jianjian, Zhang Yuanyuan
Institute of Maize and Featured Upland Crops, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310004, China.
The State Key Laboratory for Conservation and Utilization of Subtropical Agro-bioresources, South China Agricultural University, Guangzhou, 510642, China.
Biochem Biophys Res Commun. 2024 Apr 9;703:149637. doi: 10.1016/j.bbrc.2024.149637. Epub 2024 Feb 6.
The normal development of pollen grains and the completion of double fertilization in embryos are crucial for both the sexual reproduction of angiosperms and grain production. Actin depolymerizing factor (ADF) regulates growth, development, and responses to biotic and abiotic stress by binding to actin in plants. In this study, the function of the ZmADF1 gene was validated through bioinformatic analysis, subcellular localization, overexpression in maize and Arabidopsis, and knockout via CRISPR/Cas9. The amino acid sequence of ZmADF1 exhibited high conservation and a similar tertiary structure to that of ADF homologs. Subcellular localization analysis revealed that ZmADF1 is localized mainly to the nucleus and cytoplasm. The ZmADF1 gene was specifically expressed in maize pollen, and overexpression of the ZmADF1 gene decreased the number of pollen grains in the anthers of transgenic Arabidopsis plants. The germination rate of pollen and the empty seed shell rate in the fruit pods of the overexpressing plants were significantly greater than those in the wild-type (WT) plants. In maize, the pollen viability of the knockout lines was significantly greater than that of both the WT and the overexpressing lines. Our results confirmed that the ZmADF1 gene was specifically expressed in pollen and negatively regulated pollen quantity, vigor, germination rate, and seed setting rate. This study provides insights into ADF gene function and possible pathways for improving high-yield maize breeding.
花粉粒的正常发育以及胚中双受精的完成对于被子植物的有性繁殖和谷物产量都至关重要。肌动蛋白解聚因子（ADF）通过与植物中的肌动蛋白结合来调节生长、发育以及对生物和非生物胁迫的反应。在本研究中，通过生物信息学分析、亚细胞定位、在玉米和拟南芥中的过表达以及利用CRISPR/Cas9进行敲除，验证了ZmADF1基因的功能。ZmADF1的氨基酸序列表现出高度保守性，并且其三级结构与ADF同源物相似。亚细胞定位分析表明，ZmADF1主要定位于细胞核和细胞质。ZmADF1基因在玉米花粉中特异性表达，并且ZmADF1基因的过表达降低了转基因拟南芥植株花药中的花粉粒数量。过表达植株花粉的萌发率和果荚中的空籽壳率显著高于野生型（WT）植株。在玉米中，敲除系的花粉活力显著高于WT系和过表达系。我们的结果证实，ZmADF1基因在花粉中特异性表达，并对花粉数量、活力、萌发率和结实率起负调控作用。本研究为ADF基因功能以及提高高产玉米育种的可能途径提供了见解。
