Inhibition of ELK1 promotes osteogenic differentiation of periodontal ligament stem cells by reducing ubiquitin-mediated degradation of DLX2.

Periodontal ligament stem cells (PDLSCs) are promising for oral and maxillofacial bone regeneration, but the mechanisms regulating their osteogenic differentiation remain unclear. This study aimed to determine the role of DLX2 and its regulation by ELK1, STUB1, and USP11 in PDLSCs. PDLSCs were genetically manipulated using lentiviral vectors. Osteogenesis was assessed by ALP and Alizarin Red S staining and expression of osteogenic markers. Protein interactions and ubiquitination were examined by co-immunoprecipitation and cycloheximide-chase assays. ELK1-dependent STUB1 transcription was verified by ChIP and luciferase reporter assays. A mouse mandibular defect model provided in vivo validation. DLX2 overexpression increased ALP activity, mineralization, and RUNX2, OSX, and OCN expression, whereas DLX2 silencing had opposite effects. DLX2-overexpressing PDLSCs enhanced bone formation in mandibular defects. STUB1 interacted with DLX2 and promoted its ubiquitination and degradation, whereas STUB1 knockdown stabilized DLX2. USP11 bound DLX2, reduced its ubiquitination, and stabilized the protein. ELK1 bound and activated the STUB1 promoter. ELK1 silencing reduced STUB1 expression, increased DLX2 and osteogenic markers, and enhanced bone formation; STUB1 overexpression partially reversed these effects. ELK1 suppresses PDLSC osteogenesis through STUB1-mediated DLX2 degradation, while USP11 stabilizes DLX2. This ELK1-STUB1-DLX2 axis may be a potential target for craniofacial bone regeneration.