The GhANT-GoPGF module regulates pigment gland development in cotton leaves.

Gossypium spp. pigment glands are a good model for studying plant secretory cavity structures. GoPGF (GOSSYPIUM PIGMENT GLAND FORMATION) is a well-characterized master transcription factor that controls gland formation in cotton; however, little is known about its transcriptional regulation. This study integrates yeast one-hybrid sequencing data and the previously reported single-cell RNA sequencing data to identify upstream GoPGF binding proteins. Several transcription factors preferentially expressed in pigment gland cells (PGCs) are identified, including the cotton AINTEGUMENTA ortholog GhANT. Silencing of GhANT produces a defective leaf-specific PGC phenotype. Knockdown of GhANT reduces mesophyll gland number and gossypol production, while CRISPR-mediated GhANT knockout suppresses mesophyll development. Overexpression of GhANT increases organ size but not cell size. GhANT binds to two CCG boxes in the GoPGF promoter to trigger GoPGF-GhJUB1-regulated gland formation. Our study dissects the subtle regulation of tissue-specific gland morphogenesis in cotton and provides molecular mechanisms to study secretory cavity structures widespread among vascular plants.