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Identification of hub genes and stepwise regulation of the polyphenol biosynthesis pathway in walnut thin skin using co-expression network analysis and RT-qPCR
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Hanieh Shahraki Jazinaki   , Baratali Fakheri * , Nafiseh Mahdinezhad , Maryam Allahdou |
| University of Zabo , fakheri@uoz.ac.ir |
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Abstract: (264 Views) |
Identification of Hub Genes and Stage-Specific Regulatory Patterns of the Polyphenol Biosynthesis Pathway in Walnut Pellicle Using Co-Expression Network Analysis and RT-qPCR. Polyphenols are major secondary metabolites in walnut and play a crucial role in the nutritional quality and antioxidant properties of the walnut pellicle. Despite their importance, the stage-specific regulation and key regulatory genes involved in the polyphenol biosynthesis pathway during walnut pellicle development remain poorly understood. In this study, the expression patterns of ten key genes involved in polyphenol biosynthesis, including PAL, C4H, 4CL, CHS, CHI, F3H, DFR, ANS, ANR, and LAR, were investigated in walnut pellicle across different developmental stages (8 to 19 weeks after flowering) using co-expression network analysis and RT-qPCR. Heatmap and correlation analyses revealed that week 9 after flowering represents a critical stage with the highest transcriptional activity of the polyphenol biosynthesis pathway, characterized by significant upregulation of most downstream genes. Co-expression network analysis identified CHS, CHI, F3H, DFR, ANR, C4H, and 4CL as hub genes with strong positive correlations, whereas PAL exhibited strong negative correlations with downstream genes, suggesting a potential feedback regulatory role in the pathway. RT-qPCR results confirmed the significant upregulation of hub genes at week 9 and showed strong consistency with the bioinformatic and network-based analyses. Overall, this study is the first to demonstrate that week 9 is a key regulatory stage in walnut pellicle development and to identify major hub genes governing polyphenol biosynthesis. These findings provide valuable insights into the regulatory mechanisms underlying polyphenol production and offer potential genetic targets for improving phenolic content and quality in walnut.
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Article number: 7 |
| Keywords: Walnut pellicle, Polyphenol biosynthesis, Flavonoid pathway, Hub genes, Co-expression network analysis, RT-qPCR |
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Type of Study: Research |
Subject:
Plant
Received: 2026/02/20 | Accepted: 2026/06/22 | Published: 2026/06/22
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