The primary cell wall is a fundamental plant constituent that is flexible but sufficiently rigid to support the plant cell shape. Although many studies have demonstrated that reactive oxygen species (ROS) serve as important signaling messengers to modify the cell wall structure and affect cellular growth, the regulatory mechanism underlying the spatial-temporal regulation of ROS activity for cell wall maintenance remains largely unclear. Here, we demonstrate the role of the Arabidopsis (Arabidopsis thaliana) multicopper oxidase-like protein skewed 5 (SKU5) and its homolog SKU5-similar 1 (SKS1) in root cell wall formation through modulating ROS homeostasis. Loss of SKU5 and SKS1 function resulted in aberrant division planes, protruding cell walls, ectopic deposition of iron, and reduced nicotinamide adeninedinucleotide phosphate (NADPH) oxidase-dependent ROS overproduction in the root epidermis–cortex and cortex–endodermis junctions. A decrease in ROS level or inhibition of NADPH oxidase activity rescued the cell wall defects of sku5 sks1 double mutants. SKU5 and SKS1 proteins were activated by iron treatment, and iron over-accumulated in the walls between the root epidermis and cortex cell layers of sku5 sks1. The glycosylphosphatidylinositol-anchored motif was crucial for membrane association and functionality of SKU5 and SKS1. Overall, our results identified SKU5 and SKS1 as regulators of ROS at the cell surface for regulation of cell wall structure and root cell growth.

Stone cells are a remarkable trait of pear (Pyrus spp.) fruit, with their abundance directly affecting fruit quality and varying significantly among cultivars. While plant hormones influence fruit development, their role in stone cell formation remains poorly understood. Here, we conducted a co-expression network analysis using RNA-seq data from 206 pear (Pyrus pyrifolia), linking hormone biosynthesis genes with known regulators of stone cell formation. This analysis identified that gibberellin (GA) 2-oxidase 1-1 (GA2ox1-1), a GA oxidase gene whose expression correlates significantly with stone cell content across cultivars, increases during key developmental stages, and localizes specifically to stone cells, as revealed by RNA in situ hybridization. Functional validation showed that expression of GA2ox1-1 in pear fruit, callus, and Arabidopsis (Arabidopsis thaliana) resulted in significantly lower lignin content and markedly lower active GA levels. To elucidate the GA-mediated regulation of stone cell lignification, we identified the DELLA protein REPRESSOR of ga1-3 Like 3 (RGL3), a key component of the GA signaling pathway. RGL3 interacts with myeloblastosis transcription factor MYB169, a known positive regulator of lignin biosynthesis, and suppresses MYB169-mediated activation of lignin biosynthesis genes, thereby negatively regulating lignin biosynthesis. However, in the presence of GA, RGL3 inhibition was relieved, leading to MYB169 activation, upregulation of lignin biosynthesis genes, and enhanced lignin deposition. These findings clarify the role of GA in stone cell formation and reveal a regulatory mechanism involving the RGL3–MYB169 module. This work provides insights into modulating stone cell content to improve pear fruit quality.

Long non‐coding RNAs (lncRNAs) play essential roles in plant growth and development. However, their expression variability and contribution to phenotypic changes during the domestication of fruit crops remain unclear. Here, we generated approximately 3.5T of strand‐specific RNA sequencing data from wild, landrace, and improved pear varieties and identified 12 422 lncRNAs that exhibited higher tissue specificity and lower sequence conservation than protein‐coding genes. Roughly 74% of lncRNAs overlapped with transposable elements (TEs) and significantly impacted sequence diversity, expression variation, and tissue‐specific expression. DNA methylation explained approximately 32% of the expression variation of lncRNAs across accessions. Selective sweeps revealed 857 and 519 lncRNAs under domestication and improvement, respectively. These selected lncRNAs were associated with traits such as fruit size, sugar content, and stone cell content. Metabolome profiling revealed a consistent reduction in lignin‐related metabolites during pear domestication and improvement. Integrating transcriptome and metabolome analyses, we established a lncRNA‐mRNA co‐expression network associated with lignin biosynthesis. The regulatory role of lncRNA‐pys within this network was functionally validated through transgenic experiments, demonstrating that overexpressing lncRNA‐pys led to an increase in lignin content in pear. This study represents the first effort to unveil the contributions of lncRNAs under human selection to the domestication traits of fruit crops.