PuHox52-mediated hierarchical multilayered gene regulatory network promotes adventitious root formation inPopulus ussuriensis

Adventitious root (AR) formation is critically important in vegetative propagation through cuttings in some plants, especially woody species. However, the underlying molecular mechanisms remain elusive. Here, we report the identification of a poplar homeobox gene,PuHox52, which was induced rapidly (within 15 min) at the basal ends of stems upon cutting and played a key regulatory role in adventitious rooting. We demonstrated that overexpression ofPuHox52significantly increased the number of ARs while suppression ofPuHox52had the opposite effect. A multilayered hierarchical gene regulatory network (ML-hGRN) mediated by PuHox52 was reverse-engineered and demonstrated to govern AR formation. PuHox52 regulated AR formation through upregulation of nine hub regulators, including a jasmonate signaling pathway gene,PuMYC2, and an auxin signaling pathway gene,PuAGL12. We also identified coherent type 4 feed-forward loops within this ML-hGRN; PuHox52 repressedPuHDA9, which encodes a histone deacetylase, and led to an increase in acetylation and presumably expression of three hub regulators,PuWRKY51,PuLBD21andPuIAA7. Our results indicate that the ML-hGRN mediated by PuHox52 governs AR formation at the basal ends of stem cuttings from poplar trees.