Wood formation involves consecutive developmental steps, including cell division of vascular cambium, xylem cell expansion, secondary cell wall (SCW) deposition, and programmed cell death. In this study, we identified PagMYB31 as a coordinator regulating these processes in Populus alba × Populus glandulosa and built a PagMYB31-mediated transcriptional regulatory network. PagMYB31 mutation caused fewer layers of cambial cells, larger fusiform initials, ray initials, vessels, fiber and ray cells, and enhanced xylem cell SCW thickening, showing that PagMYB31 positively regulates cambial cell proliferation and negatively regulates xylem cell expansion and SCW biosynthesis. PagMYB31 repressed xylem cell expansion and SCW thickening through directly inhibiting wall-modifying enzyme genes and the transcription factor genes that activate the whole SCW biosynthetic program, respectively. In cambium, PagMYB31 could promote cambial activity through TRACHEARY ELEMENT DIFFERENTIATION INHIBITORY FACTOR (TDIF)/PHLOEM INTERCALATED WITH XYLEM (PXY) signaling by directly regulating CLAVATA3/ESR-RELATED (CLE) genes, and it could also directly activate WUSCHEL HOMEOBOX RELATED4 (PagWOX4), forming a feedforward regulation. We also observed that PagMYB31 could either promote cell proliferation through the MYB31-MYB72-WOX4 module or inhibit cambial activity through the MYB31-MYB72-VASCULAR CAMBIUM-RELATED MADS2 (VCM2)/PIN-FORMED5 (PIN5) modules, suggesting its role in maintaining the homeostasis of vascular cambium. PagMYB31 could be a potential target to manipulate different developmental stages of wood formation.

Maintenance of intracellular ion balance, especially Na⁺ and K⁺, plays an important role in plant responses to salt stress. Vessels in xylem are responsible for long-distance ion transport in vascular plants. Knowledge on the salt stress response in woody plants in limited. In this study, we identified miR319a as an important regulator in respond to salt stress in poplar. miR319a overexpression transgenic poplar showed a salt-tolerant phenotype, and cytological observation showed reduced cambium cell layers, wider xylem, increased number and lumen area of vessels and fibers, and thinner cell wall thickness in the transgenics. The miR319a-MIMIC plants, meanwhile, had opposite phenotypes, with narrower xylem, reduced number and lumen area of vessels and fibers cells, and increased wall thickness. In addition, overexpression of miR319a driven by the vessel-specific promoter significantly improved the salt tolerance compared with the fiber-specific promoter. The expression levels of PagHKT1;2 and PagSKOR1-b, which encoded high-affinity K⁺ and Na⁺ transporters for Na⁺ efflux and K⁺ influx, respectively, were positively correlated with the vessel number and lumen area. These results suggest that miR319 not only promotes ion transport rates by increasing vessel number and lumen area and reducing cell wall thickness, but also regulates the concentrations of Na⁺ and K⁺ in the xylem by up-regulating PagHKT1;2 and PagSKOR1-b. We demonstrate that miR319 may coordinate the response of poplar to salt stress through both mechanisms, enriching our understanding of the synergistic effects of the secondary xylem structure and long-distance ion transport balance in the salt tolerance of poplar.

The Jumonji C (JMJ-C) domain-containing gene family regulates epigenetic and developmental processes in plants. We identified 55 JMJ-C genes in Populus alba × Populus glandulosa using HMM and BLASTp analyses. Chromosomal mapping revealed an asymmetric distribution with conserved synteny. Phylogenetic reconstruction revealed that PagJMJ genes segregate into five evolutionarily conserved subfamilies, exhibiting classification patterns identical to those of Arabidopsis thaliana and Populus trichocarpa. Synteny analysis indicated a closer relationship with P. trichocarpa than with A. thaliana. Motif and promoter analyses highlighted subfamily-specific features and diverse cis-elements, particularly light-responsive motifs. Expression profiling revealed tissue-specific patterns, with key genes enriched in roots, vascular tissues, and leaves. Developmental analysis in cambium and xylem identified four expression clusters related to wood formation. Co-expression analysis identified six key PagJMJ genes (PagJMJ6, 29, 34, 39, 53, and 55) strongly associated with wood formation-related transcription factors. ChIP-qPCR analysis revealed that key genes co-expressed with PagJMJ genes were marked by H3K4me3 and H3K9me2 modifications. These findings provide insights into the evolutionary and functional roles of PagJMJ genes in poplar vascular development and wood formation.