TargetGAN: A generative AI framework for the design of plant core promoters with targeted activity

Plant core promoters (PCPs) are key genetic elements controlling gene expression, holding significant value for crop breeding and plant synthetic biology. However, natural promoters (NPs) are constrained by limited diversity and a narrow activity range, and it remains unclear whether synthetic promoters (SPs) can transcend these natural constraints. Here, we present TargetGAN, a deep learning framework trained on 76,851 NPs that integrates generative adversarial networks (GANs) with a pre-trained activity predictor to enable the de novo design of PCPs with user-defined activity. We used TargetGAN to generate 55,296 SPs, selecting 5,250 for high-throughput functional validation using STARR-seq. Of these, 2,909 were successfully characterized, exhibiting a moderate correlation (PCC = 0.6435) between predicted and experimental activity. Surprisingly, 29 of these SPs exhibited ultra-high activity, exceeding the maximum activity of tested NPs. Further orthogonal validation via luciferase (LUC) reporter assays demonstrated a strong positive correlation with STARR-seq measurements across a broad dynamic range. Notably, the most active synthetic candidate, SP1482, significantly outperformed the strongest tested NP, the UBI-core-promoter, achieving a 128-fold expression increase relative to the 35S minimal promoter. Interpretable motif analysis suggests that ultra-high-activity promoter design can be achieved through the precise arrangement of strong activating motifs. These results demonstrate that TargetGAN is a robust and generalizable framework for the targeted generation of PCPs tailored to user-defined targets, and will be a powerful tool both for precise gene regulation in plant systems and for overexpression analysis in genetic engineering and synthetic biology.