Jul 2026· Plant, Cell and Environment· 0 citations· 91 references
Medicine
TL;DR
A WRKY-MYB cascade that mediates SCW formation in H. brasiliensis is revealed, extending the classical NAC-MYB regulatory framework and providing molecular targets for improving xylem properties and stress resilience in tropical perennial crops.
Abstract
Secondary xylem differentiation determines wood structure and function in perennial plants, yet its regulatory mechanisms remain poorly understood in Hevea brasiliensis. Here, we generated a single-nucleus RNA sequencing (snRNA-seq) atlas from the cambium to mature xylem region in stems, resolving transcriptionally distinct cell populations corresponding to cambium, xylem mother cells, and fibre-vessel cells. Pseudotime analysis reconstructed the developmental trajectories from cambium to late xylem cell states and revealed stage-specific activation of secondary cell wall (SCW) biosynthesis programmes. Gene regulatory network analysis identified HbWRKY12a as a fibre-vessel enriched transcription factor functioning as a key regulator in SCW formation. Molecular assays demonstrated that HbWRKY12a directly binds and activates HbMYB1R1c, which subsequently modulates lignin-associated SCW deposition. Heterologous overexpression of either HbWRKY12a or HbMYB1R1c in Arabidopsis resulted in significant reductions in stem diameter and plant height. Both overexpression lines exhibited markedly decreased SCW thickness in fibre, vessels, and pith cells. These findings reveal a WRKY-MYB cascade that mediates SCW formation in H. brasiliensis, extending the classical NAC-MYB regulatory framework and providing molecular targets for improving xylem properties and stress resilience in tropical perennial crops.
Rubber tree is an economically important woody plant, whereas the cellular heterogeneity and regulatory networks underlying its xylem differentiation remain poorly characterised. Here, we built a protocol to isolate differentiating xylem protoplasts and performed scRNA-seq on two rubber tree cultivars (RK525 and RY73397) with distinct rates of girth growth. The first single-cell transcriptional atlas of differentiating xylem in the rubber tree was established, providing a cellular basis for growth variations. Ten distinct cell clusters were identified, with cell types-including vessel element, libriform fibre, and ray parenchyma-determined by established marker genes. Pseudotime analysis revealed two clearly developmental trajectories, underscoring fusiform lineages (fibres and vessels) being crucial for growth differences. Specifically, the fast-growing cultivar RK525 exhibited a cluster of upregulated genes in fibre and vessel cells. Subsequently, differentially expressed genes (DEGs) in fusiform cell clusters between the two cultivars were intersected with subcluster-upregulated genes and pseudotime-core DEGs in RK525, highlighting secondary cell wall (SCW) biosynthesis genes that regulate xylem development and promote rapid growth. WGCNA further identified a specific module significantly correlated with fibre cells, which includes six key transcription factors. Moreover, ATAC-seq revealed variety-specific chromatin accessibility patterns and transcription factor binding motifs associated with xylem development. Functional validation using Arabidopsis mutants of rubber tree homologues confirmed their crucial roles in vascular development. These findings reveal intricate regulatory programmes in differentiating xylem, and provide valuable resources and targets for genetic improvement of the rubber tree.
Chaofeng Zhang, Yuan Yao, Miaomiao Zhou et al.· Plant, Cell and Environment· 0 citations
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.
Conifers exhibit distinct age-related transitions in wood development, yet the molecular mechanisms governing these changes remain poorly characterized. Here, we integrate multi-omics and functional analyses to unravel a transcriptional regulatory network underlying age-related wood formation in Chinese pine (Pinus tabuliformis). Cross-sectional and Raman microspectroscopy analyses revealed age-dependent enhancement of lignification and carbohydrate deposition in secondary xylem, coinciding with increased pith proportion. Temporal transcriptome profiling identified PtARN (AGE-RELATED NAC TRANSCRIPTION FACTOR) as a key age-correlated regulator, whose expression is directly activated by the conserved age timer PtDAL1 through promoter binding. We conducted a genome-wide identification of secondary cell wall biosynthesis pathway genes in P. tabuliformis, and genome-wide DNA affinity purification analysis uncovered that PtARN direct binding to promoters of 189 secondary cell wall biosynthesis genes. Strikingly, PtARN represses PtMYB162, a lignin biosynthesis suppressor, forming a regulatory cascade to fine-tune lignification. Collectively, our work establishes PtARN as a master regulator orchestrating age-dependent secondary cell wall deposition and lignification in the secondary xylem of Chinese pine, and delineates a previously uncharacterized PtDAL1-PtARN-mediated molecular regulatory network in P. tabuliformis. These findings significantly advance our understanding of the molecular regulatory network underlying wood formation in conifers, and bridging developmental timing with cell wall remodelling, and provide a robust framework for conifer-specific wood formation studies.
Quan Zuo, Qian Sun, Pei-Yi Wang et al.· Plant, Cell and Environment· 0 citations
Xylem development involves a series of coordinated processes, including cell differentiation, expansion, secondary cell wall (SCW) deposition, and programmed cell death (PCD). Here, we characterize a subtilisin-like serine protease gene, SBT4.1, which exhibits specific expression in stem xylem. Loss of SBT4.1 function results in reduced xylem cell number, smaller cell size, thinner secondary walls, and delayed organelle degradation during PCD. In contrast, SBT4.1 overexpression accelerates cellular clearance and increases xylem cell number, size, and wall thickness, indicating its critical roles in differentiation, SCW synthesis, and PCD. Consistent with a positive regulatory function, pectinase activity and the expression of pectin-related genes are decreased in the mutant but increased in overexpressors. Accordingly, SBT4.1 promotes cell expansion, at least partly, by modulating pectinase activity, as evidenced by enhanced pectin methylesterase (PME) activity upon co-expression with specific PME genes in Nicotiana benthamiana. Molecular analyses revealed that SBT4.1 is a direct transcriptional target of the key SCW regulators SND1, VND6, and MYB46. Furthermore, the expression of SCW synthesis-related genes and PCD-associated protease genes is downregulated in the mutant and upregulated in overexpressing plants. Taken together, our findings demonstrate that SBT4.1, transcriptionally activated by SND1/VND6/MYB46, coordinates xylem cell differentiation, SCW deposition, and PCD, while also promoting cell expansion through modulation of pectin metabolism.
Chong Zhang, Yaoming Xu, Ziyu Ge et al.· Plant physiology and biochem...· 0 citations
Stem growth and development greatly influence on plant architecture and yield, which are intimately associated with cell wall remodeling. However, the regulatory mechanisms governing cell-wall remodeling in crops remain poorly understood. Xyloglucan endotransglucosylase/hydrolase (XTH) plays a pivotal role in xyloglucan metabolism. Here, we demonstrated that SlXTH19 controls cell wall remodeling and stem growth. Knockout or knockdown of SlXTH19 decreased cell wall thickness and cellulose and hemicellulose content in tomato stems. We demonstrated that ETHYLENE RESPONSE FACTOR (ERF) transcription factor SlERF1B binds directly to the SlXTH19 promoter, repressing its expression. Knockdown of SlERF1B showed greater stem growth and cell wall composition, whereas SlERF1B-overexpressing plants had a shorter plant heights and thinner stems than the wild type. Further investigation revealed that SlERF1B directly binds to the SlGA3ox1 promoter and represses production of GA. In addition, SlGAI, the core negative regulator of GA signaling, interacts with SlERF1B, and these interactions enhanced the inhibitory effect of SlERF1B on its downstream targets SlXTH19 and SlGA3ox1. Collectively, these results reveal a signaling module in which GA signaling modulates the degradation of SlGAI, attenuates ethylene (ETH) signaling to inhibit SlXTH19 expression, and regulates stem growth in tomato plants.
Junfeng Luo, Xi Wang, Qiongqiong Zhang et al.· Plant Physiology· 0 citations
Auxin Response Factors (ARFs) are of vital importance in plant growth and vascular development. Class A ARFs are the core auxin-driven drivers of embryogenesis, vascular pattern development, (pro)cambial stem cell initiation and xylem cell fate specification, while class B ARFs may attenuate or fine-tune auxin-mediated development. The potential roles of class C ARFs in vascular development and xylogenesis, however, remain largely unexplored. In this study, we identified Eucalyptus grandis ARF10 (EgrARF10) as a potential regulator of secondary cell wall (SCW) development. We show that EgrARF10 is nucleus-localised and strongly associated with SCW biosynthetic genes and transcription factors across co-expression and multi-omic networks. Mining of published spatial and single cell transcriptomic data revealed preferential expression of EgrARF10 in vessel and fusiform organizer cells of secondary xylem tissue, while EgrARF10 orthologs in other species exhibit diverse cell type-specific expression ranging from root cap vascular and metaxylem to cork cambium. Heterologous overexpression of EgrARF10 in hybrid poplar did not alter overall growth or morphology but resulted in a significant reduction in stem lignification, accompanied by relative increases in D-glucose, D-xylose and D-mannose, indicating altered SCW composition. While the molecular mechanism by which EgrARF10 acts remains unknown, these findings provide the first evidence for the role of a class C ARF in xylem SCW biology.
Ipeleng Makhura, R. Ployet, A. Myburg et al.· Tree Genetics & Genomes· 0 citations