Editing melon eIF4E associates with virus resistance and male sterility,Plant Biotechnology Journal

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Editing melon eIF4E associates with virus resistance and male sterility
Plant Biotechnology Journal ( IF 10.1 ) Pub Date : 2022-07-01 , DOI: 10.1111/pbi.13885
Giuliano S Pechar ₁ , Livia Donaire ₁ , Blanca Gosalvez ₁ , Carlos García-Almodovar ₁ , María Amelia Sánchez-Pina ₁ , Verónica Truniger ₁ , Miguel A Aranda ₁

Affiliation

The cap-binding protein eIF4E, through its interaction with eIF4G, constitutes the core of the eIF4F complex, which plays a key role in the circularization of mRNAs and their subsequent cap-dependent translation. In addition to its fundamental role in mRNA translation initiation, other functions have been described or suggested for eIF4E, including acting as a proviral factor and participating in sexual development. We used CRISPR/Cas9 genome editing to generate melon eif4e knockout mutant lines. Editing worked efficiently in melon, as we obtained transformed plants with a single-nucleotide deletion in homozygosis in the first eIF4E exon already in a T0 generation. Edited and non-transgenic plants of a segregating F2 generation were inoculated with Moroccan watermelon mosaic virus (MWMV); homozygous mutant plants showed virus resistance, while heterozygous and non-mutant plants were infected, in agreement with our previous results with plants silenced in eIF4E. Interestingly, all homozygous edited plants of the T0 and F2 generations showed a male sterility phenotype, while crossing with wild-type plants restored fertility, displaying a perfect correlation between the segregation of the male sterility phenotype and the segregation of the eif4e mutation. Morphological comparative analysis of melon male flowers along consecutive developmental stages showed postmeiotic abnormal development for both microsporocytes and tapetum, with clear differences in the timing of tapetum degradation in the mutant versus wild-type. An RNA-Seq analysis identified critical genes in pollen development that were down-regulated in flowers of eif4e/eif4e plants, and suggested that eIF4E-specific mRNA translation initiation is a limiting factor for male gametes formation in melon.

中文翻译：

编辑甜瓜 eIF4E 与病毒抗性和雄性不育相关

帽结合蛋白 eIF4E 通过与 eIF4G 相互作用，构成 eIF4F 复合物的核心，在 mRNA 的环化及其随后的帽子依赖性翻译中发挥关键作用。除了在 mRNA 翻译起始中的基本作用外，eIF4E 还具有其他功能，包括充当前病毒因子和参与性发育。我们使用 CRISPR/Cas9 基因组编辑来生成甜瓜eif4e敲除突变株系。编辑在甜瓜中有效地进行，因为我们获得了 T0 代中第一个eIF4E外显子纯合性单核苷酸缺失的转化植物。用摩洛哥西瓜花叶病毒（MWMV）接种分离 F2 代的编辑和非转基因植物；纯合突变植物表现出病毒抗性，而杂合和非突变植物则被感染，这与我们之前在eIF4E中沉默植物的结果一致。有趣的是，T0和F2代的所有纯合编辑植物均表现出雄性不育表型，而与野生型植物杂交则恢复了育性，显示出雄性不育表型的分离与eif4e突变的分离之间的完美相关性。对甜瓜雄花在连续发育阶段的形态学比较分析表明，小孢子母细胞和绒毡层均在减数分裂后发育异常，突变体与野生型的绒毡层降解时间存在明显差异。 RNA-Seq 分析鉴定了花粉发育中的关键基因，这些基因在eif4e/eif4e植物的花朵中下调，并表明 eIF4E 特异性 mRNA 翻译起始是甜瓜雄配子形成的限制因素。

更新日期：2022-07-01

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