An InDel in the PmARF9 promoter regulates fruit shape in Prunus mume via allele-specific binding of PmKAN1

• The genome-wide association study of 5 fruit morphology-related traits, providing data support for dissecting the genetic mechanism of fruit morphology in Prunus mume . • An InDel variation (AG-del/AG-ins) at the 372 bp position in the promoter of the auxin response factor gene PmARF9 is the core locus regulating fruit shape. • Overexpression experiments confirmed that PmARF9 affects fruit morphology by regulating hormone signaling and the expression of genes related to cell division and expansion. • PmKAN1 regulates fruit shape by specifically binding to the promoter of PmARF9 AG-ins and inhibiting its expression. Fruit morphology is important trait for processing in Prunus mume . However, the genetic and molecular mechanisms underlying the variation in P. mume remain unclear. To fill this gap, this study performed whole-genome resequencing on 153 P mume germplasms and conducted a genome-wide association study for five fruit morphology-related traits. We identified PmARF9 , an auxin response factor gene, as a key candidate associated with natural variation in fruit morphology. A promoter InDel located 372 bp upstream of the start codon formed two haplotypes, AG-ins and AG-del, and accessions carrying the AG-del allele exhibited significantly larger fruit transverse/longitudinal diameters and higher single-fruit weight compared to those carrying the AG-ins allele. Heterologous overexpression experiments of PmARF9 in tomato revealed that PmARF9 regulates fruit morphology by controlling hormone signaling pathways in the early stage of fruit development and the expression of genes related to cell division/expansion. Electrophoretic mobility shift assays and dual-luciferase analyses further demonstrated that PmKAN1 specifically binds to the AG-ins promoter haplotype and represses PmARF9 transcription, but does not bind to the AG-del haplotype. Collectively, these results demonstrate that the promoter InDel alters allele-specific PmKAN1 binding, thereby differentially regulating PmARF9 expression and ultimately contributing to fruit morphology variation in P. mume .