A Novel CsYABBY3‐CsAS1 Feedback Loop Coordinates Trichome Differentiation and Cannabinoid Biosynthesis in Cannabis sativa L.

ABSTRACT Glandular trichome development typically coincides with specialized‐metabolite production, yet the regulatory mechanisms that synchronizes these processes remain elusive. We uncover a trichome‐enriched CsYABBY3‐CsAS1 transcriptional module that links glandular trichome differentiation with cannabinoid biosynthesis in cannabis. CsYABBY3, a FILAMENTOUS FLOWER (FIL)/YAB3‐family transcription factor, promotes glandular trichome formation and elevates total cannabinoid levels. It directly binds a conserved TAATTAA motif in the promoters of CsPT4 and CsCBDAS , activating their transcription. The R2R3‐MYB CsAS1 can also stimulate cannabinoid pathway gene expression independently, but its activity is enhanced in partnership with CsYABBY3. The two factors physically associate and reciprocally upregulate each other, forming a positive feedback loop that amplifies target‐gene output. A single residue (M199) in CsYABBY3 is required for CsAS1 interaction and cooperative promoter activation. Comparative evidence suggests broadly conserved FIL/YAB3‐AS1 coupling across species, alongside lineage‐specific differences in the molecular basis of complex formation. These findings define an engineerable regulatory circuit that connects trichome fate with specialized metabolism and provide a practical route to improve cannabinoid yield.