Overexpression of MtDof32 in Medicago truncatula Enhances Leaf and Flower Organ Size through Modulation of Cell Expansion

Tao Guo^1,2Huan Wang³Shumin Wang^1,2*Ling Zhou⁴Shunzhao Sui^1,2*

• ¹College of Horticulture and Landscape Architecture，Southwest University, Chongqing, China
• ²Chongqing Engineering Research Center for Floriculture, Key Laboratory of Agricultural Biosafety and Green Production of Upper Yangtze River (Ministry of Education) ,Southwest University, Chongqing, China
• ³Chongqing Institute of Geology and Mineral Resources, Chongqing, China
• ⁴Chongqing Key Laboratory of Germplasm Innovation and Utilization of Native Plants, Chongqing Landscape and Gardening Research Institute, Chongqing, China

As leaf organ size is a key determinant of plant morphology and development, elucidating the cellular and molecular mechanisms governing organ size regulation has become a central research focus in legume biology. DNA binding with one finger 32 (MtDof32), previously identified as a key regulatory factor in organ development, has been shown to influence leaf and flower organ size in Arabidopsis. Here, we demonstrated that MtDof32 overexpression in Medicago truncatula resulted in the enlargement of both leaf and flower organs. Cellular analysis revealed that MtDof32 primarily modulated organ size by controlling cell expansion. Yeast two-hybrid screening combined with bimolecular fluorescence complementation (BIFC) assays definitively established the nuclear-localized interaction between MtDof32 and MtEBP1. Additionally, we observed that MtDof32 overexpression led to delayed flowering and reduced branching in transgenic plants. Comparative analysis with wild-type plants indicated significant alterations in the expression levels of key flowering and branching regulatory genes. This study underscored the conserved role of MtDof32 in regulating organ size across leguminous species, offering valuable insights for the genetic modification and utilization of this gene in crop improvement strategies.