A mutation in the intron splice acceptor site of a GA3ox gene confers dwarf architecture in watermelon ( Citrullus lanatus L.)

Dwarf architecture is an important trait associated with plant yield, lodging resistance and labor cost. Here, we aimed to identify a gene causing dwarfism in watermelon. The ‘w106’ (dwarf) and ‘Charleston Gray’ (vine) were used as parents to construct F 1 and F 2 progeny. Dwarf architecture of ‘w106’ was mainly caused by longitudinal cell length reduction and was controlled by a single recessive gene. Whole-genome sequencing of two parents and two bulk DNAs of F 2 population localized this gene to a 2.63-Mb region on chromosome 9; this was further narrowed to a 541-kb region. Within this region, Cla015407, encoding a gibberellin 3β-hydroxylase (GA3ox), was the candidate gene. Cla015407 had a SNP mutation (G → A) in the splice acceptor site of the intron, leading to altered splicing event and generating two splicing isoforms in dwarf plants. One splicing isoform retained the intron sequences, while the other had a 13-bp deletion in the second exon of GA3ox transcript, both resulting in truncated proteins and loss of the functional Fe2OG dioxygenase domain in dwarf plants. RNA-Seq analysis indicated that expression of Cla015407 and other GA biosynthetic and metabolic genes were mostly up-regulated in the shoots of dwarf plants compared with vine plants in F 2 population. Measurement of endogenous GA levels indicated that bioactive GA 4 was significantly decreased in the shoots of dwarf plants. Moreover, the dwarf phenotype can be rescued by exogenous applications of GA 3 or GA 4+7 , with the latter having a more distinct effect than the former. Subcellular localization analyses of GA3ox proteins from two parents revealed their subcellular targeting in nucleus and cytosol. Here, a GA3ox gene controlling dwarf architecture was identified, and loss function of GA3ox leads to GA 4 reduction and dwarfism phenotype in watermelon.