Integrating proteomics and phosphoproteomics to reveal the regulatory mechanism of yellowing in postharvest cold storage of broccoli

Abstract

Yellowing represents the main form of quality degradation in postharvest broccoli. In this study, the changes in protein levels and protein phosphorylation that occur in tandem with the yellowing of broccoli were analyzed using the 4D-Label free method. Through joint analysis, our focus was directed towards six KEGG-enriched pathways associated with proteins (where no protein differences were observed, yet significant phosphorylation changes took place), namely: protein processing in endoplasmic reticulum and mRNA surveillance pathway, nucleocytoplasmic transport, spliceosome, plant hormone signal transduction, and MAPK signaling pathway-plant. Among them, SNF1-related protein kinases 2 (SnRK2) and the transcription factor ABF, ethylene insensitive protein 2 (EIN2), JA-insensitive jasmonate resistant 1 (JAR1), brassinosteroid-signaling kinase (BSK), mitogen-activated protein kinase 6-Like (MAPK6-Like), G-type lectin S-receptor-like serine/threonine protein kinase (GsSRK), receptor-like kinase1, calreticulin, nonsense transcripts UPF2, serine/arginine-rich splicing factor SR45, apoptotic chromatin condensation inducer in the nucleus-like (Acinus) may indirectly affect the yellowing of broccoli. In the photosynthesis-antenna protein pathway, chlorophyll a/b-binding proteins have emerged as key targets under the purview of multiple omics analyses. These proteins, in synergy with chlorophyll-metabolizing enzymes, exerted a combined influence on the yellowing of broccoli. Simultaneously, post-harvest broccoli undergoes several notable changes. Its cell wall structure loosens, leading to a reduction in barrier-protection capabilities. Meanwhile, the defense response was enhanced, while DNA replication and mRNA synthesis were inhibited. Additionally, the processes of nonsense mutation and cleavage of degraded transcripts were impeded. Collectively, these alterations hastened the senescence process of broccoli and were likely to be the causative factors behind its yellowing.