E3 Ubiquitin Ligase March6 Suppresses Odontoblast Differentiation and Dentinogenesis

doi:10.13701/j.cnki.kqyxyj.2025.08.004

Abstract

Abstract: Objective: To investigate the role of E3 ubiquitin ligase March6 in odontoblast differentiation and dentinogenesis. Methods: Immunohistochemistry (IHC) staining was performed on mouse tooth sections at different developmental stages to determine the spatiotemporal expression pattern of March6. March6 was knocked down in mouse dental papilla cells (mDPCs) in vitro, followed by mineralization induction. Quantitative real-time PCR, western blot, alkaline phosphatase (ALP) staining, and alizarin red S staining were subsequently performed to evaluate the effect of March6 on the odontoblastic differentiation potential of mDPCs. Prx1^cre mice were crossed with March6^flox/flox mice to generate conditional knockout (cKO) mice with mesenchyme-specific deletion of March6 in the dental tissue (March6 cKO, Prx1^cre;March6^flox/flox). Specific deletion of March6 was validated by IHC. Mandibles from postnatal day 10 (PN10) March6 cKO and littermate control mice were analyzed by hematoxylin and eosin staining and micro-computed tomography (micro-CT) to evaluate the in vivo effect of March6 on dentinogenesis. Results: March6 exhibited low expression in the dental mesenchyme of embryonic mice, while high expression was observed in odontoblasts after their differentiation. Knockdown of March6 in mDPCs significantly upregulated the mRNA and protein levels of Dmp1 and Dspp during odontoblastic differentiation. Furthermore, ALP activity and calcified nodule formation were markedly increased in March6 deficient cells. Histological analysis and micro-CT imaging revealed a significantly thicker dentin layer in March6 cKO mice compared with littermate controls. Conclusion: The E3 ubiquitin ligase March6 negatively regulates odontoblast differentiation and dentinogenesis.

Key words: March6, odontoblast differentiation, dentinogenesis, gene knockout

FENG Hao, YUAN Guohua. E3 Ubiquitin Ligase March6 Suppresses Odontoblast Differentiation and Dentinogenesis[J]. Journal of Oral Science Research, 2025, 41(8): 657-663.

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