Physiological and cellular mechanisms of the transcription factor CcLBD36 positively regulating leaf size in pepper

A genome-wide association study of leaf length in 182 pepper accessions, combined with gene annotation and p-value screening, identified LBD36 as a key candidate gene associated with leaf size. Sequence characteristics, phylogenetic relationships, chromosomal localization, and promoter cis-acting elements of CcLBDs in the genome of Capsicum chinense were analyzed. CcLBD36 was cloned from Hainan huangdenglong pepper. Real-time quantitative PCR analysis showed that CcLBD36 expression was significantly higher in large-leaf pepper varieties than in small-leaf varieties, and was positively correlated with chlorophyll a, chlorophyll b, total chlorophyll content, and net photosynthetic rate. Exogenous application of IAA, GA3, and MeJA induced its upregulation, while its expression exhibited an initial increase, followed by a decline during leaf development. Virus-induced gene silencing (VIGS) targeting CcLBD36 resulted in significantly reduced leaf area, leaf length, leaf width, and plant width, along with decreased chlorophyll a, chlorophyll b, total chlorophyll content, net photosynthetic rate, stomatal conductance, intercellular CO2 concentration, and transpiration rate. Cytological analysis revealed significantly smaller cell areas in the leaf tissues of silenced plants, while hormone assays indicated a concurrent reduction in growth-promoting hormones, and a marked increase in the inhibitory hormone MeJA in these lines. In summary, CcLBD36 positively regulates pepper leaf size.