Exploratory Biological Coordinate Analysis on 2,4-Epi-brassinolide-induced the Size Change of Leaf Angle in Tobacco Seedlings

Leaf angle (LA) is a key plant architectural trait contributing to crop yield. The chemical 2,4-epibrassinolide (EBR) is a bioactive form of brassinosteroids (BRs) and plays a significant role in influencing plant architecture, including regulating the LA. To investigate the effects of EBR on LA in tobacco seedlings, we histochemically and molecularly characterized changes in LA in response to treatment with EBR at two different concentrations, T1 (1 × 10 −8 mol/L) and T2 (1 × 10 −5 mol/L). Compared with control plants, EBR significantly increased the LA in T1 and T2 by 38.6% and 37.3%, respectively, which likely resulted from increased cell areas of 95.2% and 185%, respectively, and inhibited cell division with the number of cells per unit area experiencing 16.4% and 41.8% decreases, respectively. The relative expression levels of NtBIN2 , NtEXPA28 , and NtCYCD1 genes were consistently correlated with the size of the LA. By analyzing the expression of the auxin (IAA), gibberellin (GA), and brassinosteroid (BR) biosynthesis and signaling pathways, the coordinated relationship of these hormones were evaluated. A positive correlation between EBR concentration and the expression of IAA biosynthesis ( NtYUCCA8 ), GA biosynthesis ( NtGA20 ), BR signaling ( NtBRI1 , NtBES1 ), and IAA signaling ( NtGH3.2 and NtLC3 ) pathway related genes was observed, while a negative correlation was observed for the expression of cell cycle regulating genes ( NtCYCD2 , NtCYCD3 , and NtCYCD5 ). We also found that Cyclin, GA20, BRI1 and LC3 were more likely involved in regulating LA formation in tobacco. The findings of this study expand our knowledge of how plant architectural features such as LA are regulated at the molecular level and provides potential gene targets for genetic engineering or breeding efforts aimed at improving crop productivity and yields.