VviAGL11, the Arabidopsis SEEDSTICK homolog, has been proposed to have a causative role in grapevine stenospermocarpy. An association between a mutation in the coding sequence (CDS) and the seedless phenotype was reported, however, no working mechanisms have been demonstrated yet. We performed a deep investigation of the full VviAGL11 gene sequence in a collection of grapevine varieties belonging to several seedlessness classes that revealed three different promoter-CDS combinations. By investigating the expression of the three VviAGL11 alleles, and by evaluating their ability to activate the promoter region, we observed that VviAGL11 self-activates in a specific promoter-CDS combination manner. Furthermore, by transcriptomic analyses on ovule and developing seeds in seeded and seedless varieties and co-expression approaches, candidate VviAGL11 targets were identified and further validated through luciferase assay and in situ hybridization. We demonstrated that VviAGL11 Wild Type CDS activates Methyl jasmonate esterase and Indole-3-acetate beta-glucosyltransferase, both involved in hormone signaling and Isoflavone reductase, involved in secondary metabolism. The dominant-negative effect of the mutated CDS was also functionally ectopically validated in target induction. VviAGL11 was shown to co-localize with its targets in the outer seed coat integument, supporting its direct involvement in seed development, possibly by orchestrating the crosstalk among MeJA, auxin, and isoflavonoids synthesis. In conclusion, the VviAGL11 expression level depends on the promoter-CDS allelic combination, and this will likely affect its ability to activate important triggers of the seed coat development. The dominant-negative effect of the mutated VviAGL11 CDS on the target genes activation was molecularly validated. A new regulatory mechanism correlating VviAGL11 haplotype assortment and seedlessness class in grapevine is proposed.

中文翻译：

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VviAGL11 在种子发育过程中自我调节和靶向激素和次级代谢相关基因

VviAGL11 是拟南芥 SEEDSTICK 同源物，已被提议在葡萄藤精果中具有致病作用。据报道，编码序列（CDS）中的突变与无籽表型之间存在关联，但尚未证实其工作机制。我们对属于几个无核类别的葡萄品种集合中的完整 VviAGL11 基因序列进行了深入调查，揭示了三种不同的启动子-CDS 组合。通过研究三个 VviAGL11 等位基因的表达，并通过评估它们激活启动子区域的能力，我们观察到 VviAGL11 以特定的启动子-CDS 组合方式自我激活。此外，通过对有种子和无种子品种中胚珠和发育种子的转录组分析以及共表达方法，通过荧光素酶测定和原位杂交鉴定并进一步验证了候选 VviAGL11 靶标。我们证明了 VviAGL11 野生型 CDS 激活了茉莉酸甲酯酯酶和吲哚-3-乙酸 β-葡萄糖基转移酶，两者都参与激素信号传导和异黄酮还原酶，参与次级代谢。突变 CDS 的显性负效应也在目标诱导中进行了功能异位验证。VviAGL11 被证明与外种皮外皮中的目标共定位，支持其直接参与种子发育，可能是通过协调 MeJA、生长素和异黄酮合成之间的串扰。总之，VviAGL11 表达水平取决于启动子-CDS 等位基因组合，这可能会影响其激活种皮发育的重要触发因素的能力。突变的 VviAGL11 CDS 对靶基因激活的显性负效应在分子上得到验证。提出了一种与葡萄树中的 VviAGL11 单倍型分类和无核类相关的新调控机制。