TMT-label comparative proteomics reveals the vernalization mechanism in Wucai (Brassica campestris L.)

Abstract

To investigate the molecular basis of vernalization in Wucai [Brassica campestris L. (Syn. Brassica rapa L.) ssp. chinensis var. rosularis Tsen], we performed differential proteomic analysis using a tandem mass tags (TMT)-based approach. Proteins from shoot apices subjected to 0, 15, and 30 days of vernalization (V0, V15, and V30) were analyzed to identify differentially abundant proteins (DAPs). A total of 8066 proteins were obtained, and 507 shared DAPs were involved in both initiation and progression of vernalization. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations revealed functional enrichment in cellular processes, metabolic pathways, and translation-related activities, including photosynthesis, glucosinolate biosynthesis, and flavonoid biosynthesis. Proteomic data showed reduced abundance of photosynthesis-related proteins and upregulation of flavonoid biosynthesis during vernalization. Transcriptional validation of 24 proteins across metabolic and regulatory pathways corroborated proteomic findings, with notable peaks in genes associated with flavonoid biosynthesis at 15 days of vernalization, such as VESR1,CH13, CHS1, FHT, and FLS1. The functions of these genes in vernalization will be further analyzed.

Significance

Wucai is prone to premature bolting and flowering under cold conditions, as vernalization plays a key role in controlling flowering time in Chinese cabbage crops. However, the proteomic basis of vernalization remains poorly understood. In this study, TMT-based proteomic analysis identified DAPs associated with vernalization. Pathway enrichment analysis highlighted key DAPs and their roles in significantly enriched pathways relevant to vernalization. Notably, genes in the flavonoid biosynthesis pathway genes, including VESR1, CH13, CHS1, FHT, and FLS1, respond to vernalization. These findings offer novel insights into the molecular mechanisms underlying flowering time regulation in Wucai.