Hydroxygenkwanin exerts osteoprotective effects by regulating LRG1-PJA1-PML axis.

Osteoporosis, especially postmenopausal osteoporosis driven by estrogen deficiency, remains a major public health burden due to elevated fracture risk. Current treatments are often limited by adverse effects and cost, prompting interest in safer natural compounds. Here, we identify hydroxygenkwanin (HGK), a Daphne genkwa-derived flavonoid, as a potent osteoprotective compound with an unprecedented mechanism of action. Femoral RNA sequencing from ovariectomized (OVX) mice revealed leucine-rich alpha-2-glycoprotein 1 (LRG1) as the most differentially expressed gene following HGK treatment, suggesting a core mechanism. Bioinformatics analysis and co-immunoprecipitation assays identified PRAJA ring finger ubiquitin ligase 1 (PJA1) as the E3 ubiquitin ligase that mediates the degradation of promyelocytic leukemia protein (PML), a key osteogenic regulator. In vivo, HGK effectively attenuated OVX-induced bone loss and improved bone microarchitecture through the LRG1-PJA1-PML axis. Mechanistically, we demonstrate that HGK directly binds to LRG1 (as validated by CETSA, pull-down assays, and molecular dynamics simulations), an interaction that potentiates the association of LRG1 with the E3 ubiquitin ligase PJA1. This sequestration of PJA1 prevents its interaction with, and subsequent ubiquitination of, PML. The resulting stabilization of PML promotes osteoblast differentiation in an oxidative stress milieu. Collectively, these results reveal a critical LRG1-PJA1-PML axis in OP and establish HGK as a first-in-class candidate that leverages this pathway for therapeutic intervention.