Single-nucleus RNA sequencing reveals the characteristics of anthocyanin accumulation in epidermal and mesophyll cells of Pistacia chinensis leaves.

Abstract

Anthocyanins are important plant pigments that contribute to leaf coloration and stress adaptation, but how their biosynthesis is regulated across different leaf cell types remains unclear, especially in woody plants. Here, we combined single-nucleus RNA sequencing (snRNA-seq) and bulk RNA-seq to investigate anthocyanin-associated regulatory programs in Pistacia chinensis leaves. We found that epidermal cells followed a largely unidirectional transition toward high-anthocyanin states, whereas mesophyll cells differentiated into multiple functional subtypes, revealing substantial cellular heterogeneity. Although epidermal and mesophyll cells shared the core anthocyanin biosynthetic pathway, their upstream regulatory programs differed. Integrative analysis identified multiple transcription factors associated with anthocyanin accumulation, several of which showed distinct expression patterns between high-anthocyanin epidermal cells and anthocyanin-enriched mesophyll cells. Functional assays showed that PcMYB113a and PcMYB113b promote anthocyanin accumulation and interact with conserved MBW-complex components. Overall, this study provides cell-resolved insight into anthocyanin-associated regulatory programs in woody plant leaves.