# Comparative Phosphoproteomics of Two Flammulina filiformis Cultivars with Differential Postharvest Browning Susceptibility

## TL;DR

This study compares protein phosphorylation in two Flammulina filiformis mushroom cultivars to understand why one browns more after harvest.

## Contribution

The study reveals a distinct phosphoproteomic signature linked to postharvest browning in mushrooms.

## Key findings

- The SB cultivar showed more phosphorylation changes than the RB cultivar during storage.
- Phosphorylation differences were linked to energy metabolism and kinase activity.
- Altered phosphorylation in SB correlated with greater browning and ATP loss.

## Abstract

Protein phosphorylation modification plays a role in cells’ response to oxidative stress, a key factor leading to postharvest browning of Flammulina filiformis. However, the molecular mechanism by which protein phosphorylation contributes to postharvest browning of F. filiformis remains unclear. This study aimed to characterize the basal phosphoproteomic landscapes associated with variations in different browning phenotypes of F. filiformis. Using data-independent acquisition (DIA) mass spectrometry, we comprehensively profiled the phosphorylation dynamics in susceptible-to-browning (SB) and resistant-to-browning (RB) cultivars at harvest and after 24 h storage. We identified 84,244 phosphorylation sites on 4494 phosphoproteins, with the SB cultivar displaying more altered sites (21,195) than the RB (16,087). Functional enrichment analysis revealed that the differential phosphorylation was significantly implicated in kinases and energy metabolism pathways. Notably, the SB cultivar exhibited a more pronounced phosphorylation profile on key proteins involved in ATP synthesis and glycolysis. Protein–protein interaction (PPI) network analysis further indicated a kinase-mediated regulatory network targeting core energy metabolism components, including ATP synthase and 6-phosphofructokinase. This distinct phosphosignature in the SB cultivar correlated with its more severe browning phenotype and a sharper decline in ATP content during storage. Our findings suggest that divergent phosphorylation-mediated regulation of energy metabolism is strongly associated with the differential postharvest browning susceptibility between these two cultivars, providing a valuable molecular resource for future functional studies.