SummaryWood formation, intricately linked to the carbohydrate metabolism pathway, underpins the capacity of trees to produce renewable resources and offer vital ecosystem services. Despite their importance, the genetic regulatory mechanisms governing wood fibre properties in woody plants remain enigmatic. In this study, we identified a pivotal module comprising 158 high‐priority core genes implicated in wood formation, drawing upon tissue‐specific gene expression profiles from 22 Populus samples. Initially, we conducted a module‐based association study in a natural population of 435 Populus tomentosa, pinpointing PtoDPb1 as the key gene contributing to wood formation through the carbohydrate metabolic pathway. Overexpressing PtoDPb1 led to a 52.91% surge in cellulose content, a reduction of 14.34% in fibre length, and an increment of 38.21% in fibre width in transgenic poplar. Moreover, by integrating co‐expression patterns, RNA‐sequencing analysis, and expression quantitative trait nucleotide (eQTN) mapping, we identified a PtoDPb1‐mediated genetic module of PtoWAK106‐PtoDPb1‐PtoE2Fa‐PtoUGT74E2 responsible for fibre properties in Populus. Additionally, we discovered the two PtoDPb1 haplotypes that influenced protein interaction efficiency between PtoE2Fa‐PtoDPb1 and PtoDPb1‐PtoWAK106, respectively. The transcriptional activation activity of the PtoE2Fa‐PtoDPb1 haplotype‐1 complex on the promoter of PtoUGT74E2 surpassed that of the PtoE2Fa‐PtoDPb1 haplotype‐2 complex. Taken together, our findings provide novel insights into the regulatory mechanisms of fibre properties in Populus, orchestrated by PtoDPb1, and offer a practical module for expediting genetic breeding in woody plants via molecular design.