# Transcriptional Regulatory Networks Oscillate Seasonal Plasticity of Fruit Metabolism in Melon
## 3 Results
### Transcriptomic Analysis of Melons Grown in Spring and Autumn
Due to the difference in temperature at which 0 DAP fruits grow during the spring and autumn seasons, DEGs were also enriched in the response-to-temperature stimulus (GO: 0009266) pathway. The core network detected by 10 DAP was also enriched in cell division-related genes. As the lighting time changes, there was a situation where the spring lighting time gradually exceeds the autumn (Figure 1b), and related genes of the response-to-light stimulus (GO: 0009416) began to show significance in the comparison of the two seasons. Notably, the most prominent hub gene, the ring finger ubiquitin E3 ligase, COP1, which is related to the control of plant photoperiod perception and participated in ubiquitin-mediated protein hydrolysis [24]. It was simultaneously annotated into the response-to-light stimulus (GO: 0009416) and photosynthesis (GO: 0015979) pathways. Figure 2: Co-expression analysis of hub genes in the transcriptional regulatory networks in spring- and autumn-grown melon fruits: (**a**) The heatmap illustrates the gene clustering results achieved through the Mfuzz method based on their FPKM values. It identified 12 distinct clusters with varied expression patterns. A total of 16,525 DEGs were utilized as input. The expression profiles of genes in these 12 clusters were examined. The Mfuzz clustering was executed using the clusterData function from the R package ClusterGVis, with normalization performed using the Z-score method. (**b**) PPI network in associated with melon fruit development built based on the hub genes of 40 days after pollination grown in spring and autumn comparison. Purple dashed line marks the hub gene sets of the main enriched functional pathways. Purple shaded area is the pathway with the highest number of enrichment factors. Yellow ring represents DEGs in metabolic process pathway. Blue represents down-regulated DEGs, orange represents up-regulated DEGs. Size of each node is proportional to its value of betweenness centrality and label font size is proportional to node degree. The width of the edges is represented by the co-expression value of proteins connected to the edge. (**c**) Regulatory networks of hub genes during fruit development under spring and autumn growing conditions. The color of the outer ring represents the stages they appeared as hub genes. Green, yellow, white, red, and blue represent 0, 10, 20, 30, and 40 days after pollination, respectively. Size of each node is proportional to its value of betweenness centrality and label font size is proportional to node degree. The width of the edges is represented by the co-expression value of proteins connected to the edge. (**d**) Clustered expression patterns of hub genes during fruit development under spring and autumn growing conditions. Size of each node is proportional to its FPKM values. Interestingly, the 20 DAP network was enriched with plant-type secondary cell wall biology (GO: 0009834), cell wall biology (GO: 0042546), and cellular polysaccharide biological process (GO: 0033692), which included xylan biological process (GO: 0045492), and the lignin biological process (GO: 0009809). The annotated DEGs were down-regulated in spring. The biosynthesis of these polysaccharide components and lignin were related to the formation of secondary cell walls. Lignin is an important component of the secondary cell wall, indicating that this stage underwent a secondary thickening process, significantly enhancing structural support and gradually losing the ability of cells to expand and grow, resulting in differences in pulp texture. The biosynthesis of lignin is a downstream branch of the phenylpropanol pathway, which means that the phenylpropanol genes are involved in the transcription of growth regulation mechanisms related to the content of its derived structural polymer, lignin. The photosynthesis pathway and the response-to-light stimulus pathway exhibited significant enrichment at 30 DAP, and were mainly concentrated in the response to high light intensity (GO: 0009644).