Knockout of TaOTUB1 homoeologs in wheat promotes increased grain yield and earlier flowering

Flowering and subsequent seed development are not only the most crucial reproductive activities of plants but also key determinants of crop yield. However, the mechanisms and effective strategies for the synergistic regulation of seed size and flowering time in plants remain unclear. Here, we show that TaOTUB1, a deubiquitinase gene with three homoeologs (TaOTUB1-7A, -7B, and -7D), plays a critical role in regulating grain size and flowering time in wheat. TaOTUB1 is the most highly expressed member of the wheat OTUB family and is primarily expressed at the primordia of young reproductive organs. Overexpression of TaOTUB1 results in a decrease in grain size, grain number, and grain yield, as well as delayed flowering. Conversely, knocking out all three homoeologs of TaOTUB1 increases grain size and promotes flowering, but decreases grain number and yield. Interestingly, the knockout of TaOTUB1-7A or -7B alone can increase grain size and accelerate flowering without affecting grain number, ultimately enhancing wheat yield. TaOTUB1-7A/-7B/-7D shows additive effects on grain size regulation by delaying flowering and shortening grain development time. Mechanistic studies suggest that TaOTUB1 interacts with the flowering regulator TaRCN2 in the cytoplasm, while in the nucleus, it interacts with the grain size regulator TaSPL17. Additionally, we identified a favorable haplotype HAPI with low TaOTUB1 activity, which is associated with increased grain weight, higher grain yield, and earlier flowering. These findings highlight the critical role of TaOTUB1 activity in synergistically improving grain size, yield, and flowering time in wheat.