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Pull-down results of proteins interacting with the alkaline phosphatase G-quadruplex in MC3T3-E1 cells

ABSTRACT: G-quadruplexes (G4s) are non-canonical nucleic acid structures that participate in diverse biological processes, yet their functional involvement in endogenous regulatory events governing osteogenic differentiation remains largely unexplored. In the present study, we observed a progressive decline in global G4 abundance during osteogenesis and demonstrated that small-molecule stabilization of G4 structures markedly impairs osteogenic progression in both cellular and in vivo models. To uncover the molecular basis of this regulatory effect, we employed DNA affinity precipitation combined with liquid chromatography–tandem mass spectrometry (LC-MS/MS) to identify proteins that interact with a G4 structure embedded within the promoter region of the osteogenic master regulator alkaline phosphatase (Alpl). This unbiased proteomic screening revealed multiple candidate G4-binding proteins, among which Nucleolin (NCL) and the G4-resolving helicase Dhx36 were further validated by immunoblotting. Mechanistically, these two proteins exert opposing influences on Alpl transcriptional control: Nucleolin stabilizes the Alpl promoter G4 to repress promoter activity, whereas Dhx36 unwinds the G4 structure, thereby facilitating transcriptional activation.

ORGANISM(S): Mus Musculus

SUBMITTER: Fujun Jin

PROVIDER: PXD071168 | iProX | Tue Nov 25 00:00:00 GMT 2025

REPOSITORIES: iProX

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