Mechanistic study of AP2D in the regulation of AM symbiosis

Arbuscular mycorrhizal (AM) symbiosis is a highly conserved mutualism in which plants and fungi exchange carbon for mineral nutrients, such as phosphate, via highly branched fungal arbuscules. This research investigates the regulatory architecture and mechanism downstream of the central CCaMK–CYCLOPS calcium‐signaling module, focusing on the AP2/ERF transcription factor AP2D and its paralog AP2DL. Through promoter–GUS reporter assays, we observed that AP2D/AP2DL expression is cortex‐localized and strictly induced by AM colonization in a CCaMK–CYCLOPS-dependent manner. Dexamethasone-inducible AP2D overexpression drives coordinated upregulation of strigolactone-biosynthesis genes (GGPS, D27, CCD7, CCD8, MAX1) and related apocarotenoid loci, whereas ap2d/ap2dlike double mutants exhibit phosphate- and auxin-insensitive attenuation of these transcripts. Co-expression network analysis further identified the GRAS co-regulator DIP1 as an AP2D partner. DIP1 overexpression represses AP2D/AP2DL, CCaMK, and SL-biosynthetic genes, while dip1 loss-of-function derepresses them without compromising colonization, revealing a homeostatic brake on symbiotic signaling. Collectively, these findings address AP2D as a pivotal integrator of calcium, hormonal, and nutrient cues in AM symbiosis, and identify DIP1 as a negative feedback regulator that tempers arbuscule-promoting transcriptional output.