The PtrDJ1C gene is essential for poplar growth and early chloroplast development. Disruption of PtrDJ1C expression results in an albino leaf phenotype and increased branching. However, the underlying mechanism for the increased branching remains unknown. In this study, we employed integrated approaches to investigate the function of PtrDJ1C in the branch-increasing phenotype. Our results revealed that levels of indole-3-acetic acid (IAA), gibberellin (GA), and abscisic acid (ABA) were significantly reduced in ptrdj1c mutants, while cytokinin (CK) levels were slightly increased. Transcriptomic and proteomic analyses identified several key genes and proteins involved in hormone regulation and branching development that were differentially expressed. Specifically, the expression levels of TAA, ZEP, and GA20ox-genes involved in IAA, GA, and ABA biosynthesis-were significantly reduced in ptrdj1c, while IPT and LOG, which regulate CK synthesis, were upregulated. Moreover, immunoblot analysis further validated reduced levels of key biosynthetic enzymes for IAA, GA, and ABA, alongside increased levels of IPT and LOG enzymes. Interestingly, our findings suggest that hormone signaling pathways act in concert with the transcription factor WUSCHEL (WUS) to synergistically promote branching development. These results provide novel insight into the regulatory role of PtrDJ1C in hormone balance and its downstream effects on poplar branching.