CmaGA2ox2 is associated with dwarf plant architecture controlled by the single dominant CmaDw-1 locus in winter squash

Dwarf or semi-dwarf plant architecture characterized by reduced vine or internode length is a desirable trait for winter squash (Cucurbita maxima) production, but the regulatory mechanisms and specific genes responsible for this trait remain unidentified. In this study, we conducted map-based QTL cloning and functional characterization of a major-effect QTL, underlying the dwarf phenotype (short vine length) in the inbred line Agol. A near-isogenic line, SV015 carrying the wildtype long-stem allele in Agol background was developed. Comparative histological observations in the two NILs indicated that the dwarf phenotype is due to reduced cell elongation, and thus internode length during stem elongation. Genetic analysis in NIL-derived segregating populations confirmed that a single dominant gene, CmaDw-1, controlled the dwarf phenotype. Bulked segregant analysis and fine genetic mapping identified CmaCh03G013990 as the sole candidate gene for CmaDw-1 that encodes gibberellin 2-oxidase 2 (CmaGA2ox2), a key enzyme in gibberellin (GA) metabolism. Sequence analysis revealed multiple single nucleotide polymorphisms (SNPs) or Insert/Deletions (InDels) within the promoter and coding region of CmaGA2ox2 that are associated with reduced stem elongation in Agol. Spatiotemporal expression analysis of CmaGA2ox2 revealed its differential expression in the hypocotyls between NILs in the early development stage. Quantitation of endogenous GAs and exogenous phytohormone treatments in NILs and ectopic expression of CmaDw-1 in Arabidopsis further support the critical role of CmaGA2ox2 in stem elongation. This work provides the first molecular insights into the regulation of dwarf plant architecture in C. maxima.