Non-additive expression genes play a critical role in leaf vein ratio heterosis in Nicotiana tabacum L

Heterosis, recognized for improving crop performance, especially in the first filial (F₁) generation, remains an area of significant study in the tobacco industry. The low utilization of leaf veins in tobacco contributes to economic inefficiency and resource waste. Despite the positive impacts of heterosis on crop genetics, investigations into leaf-vein ratio heterosis in tobacco have been lacking. Understanding the mechanisms underlying negative heterosis in leaf vein ratio at the molecular level is crucial for advancing low vein ratio leaf breeding research. This study involved 12 hybrid combinations and their parental lines to explore heterosis associated with leaf vein ratios. The hybrids displayed diverse patterns of positive or negative leaf vein ratio heterosis across different developmental stages. Notably, the F₁ hybrid (G70 × Qinggeng) consistently exhibited substantial negative heterosis, reaching a maximum of -19.79% 80 days after transplanting. A comparative transcriptome analysis revealed that a significant proportion of differentially expressed genes (DEGs), approximately 39.04% and 23.73%, exhibited dominant and over-dominant expression patterns, respectively. These findings highlight the critical role of non-additive gene expression, particularly the dominance pattern, in governing leaf vein ratio heterosis. The non-additive genes, largely associated with various GO terms such as response to abiotic stimuli, galactose metabolic process, plant-type cell wall organization, auxin-activated signaling pathway, hydrolase activity, and UDP-glycosyltransferase activity, were identified. Furthermore, KEGG enrichment analysis unveiled their involvement in phenylpropanoid biosynthesis, galactose metabolism, plant hormone signal transduction, glutathione metabolism, MAPK signaling pathway, starch, and sucrose metabolism. Among the non-additive genes, we identified some genes related to leaf development, leaf size, leaf senescence, and cell wall extensibility that showed significantly lower expression in F₁ than in its parents. These results indicate that the non-additive expression of genes plays a key role in the heterosis of the leaf vein ratio in tobacco. This study marks the first exploration into the molecular mechanisms governing leaf vein ratio heterosis at the transcriptome level. These findings significantly contribute to understanding leaf vein ratios in tobacco breeding strategies.