Differences in lignification between hulled and hull-less Cucurbita pepo and functional identification of the key...

The Cucurbita pepo CpMYB52 gene is a key regulator of lignin biosynthesis. Its high expression in seed coats is crucial for their development, explaining the hull-less variety's collapsed seed coat. In hull-less Cucurbita pepo, the collapse of sclerenchyma tissue leads to the disappearance of the seed coat during development. The mechanism of pericarp formation in hull-less C. pepo is not yet fully understood. Previous transcriptome results indicated that R2R3-MYB transcription factors can regulate the biosynthesis of lignin. This study identified the R2R3-MYB transcription factor family in C. pepo based on whole-genome analysis and conducted cloning and real-time quantitative fluorescence research on the CpMYB52. The results showed that there are a total of 186 R2R3-MYB transcription factor members in C. pepo, distributed across 20 chromosomes. Phylogenetic analysis divided the R2R3-MYB gene family of C. pepo into 29 subgroups (C1 ~ C29), with differences in gene structure among members of different subgroups. Cloning analysis of the CpMYB52 gene showed that the gene encodes a peptide with 187 amino acids. Phylogenetic analysis showed that the CpMYB52 protein had the strongest speciation specificity and the closest ties to the C. moschata and C. maxima species. Temporal expression analysis revealed that CpMYB52 is specifically expressed during seed coat development in normal C. pepo, at levels 30 times higher than those observed in hull-less C. pepo during the same period. Furthermore, the analysis of tissue-specific expression revealed a significant tissue specificity of the CpMYB52 gene, exhibiting varying expression levels in different tissues. The expression pattern of this gene was higher in more lignified sites, such as roots and stems. In both pumpkin varieties, CpMYB52 was expressed 15-fold higher in roots and 7.7-fold higher in stems than in cotyledons. In Arabidopsis, overexpression of CpMYB52 enhances S-lignin synthesis and increases the total amount of lignin. The expression of AtC3H, At4CL, and AtF5H genes in transgenic Arabidopsis was 134-fold, 48-fold, and 22-fold higher than in wild-type Arabidopsis. The CpMYB52 gene can positively regulate lignin biosynthesis and change the structure of the lignin monomer. These results suggest that the CpMYB52 gene plays a crucial role in regulating seed coat development and morphogenesis.