LAUSR

N6‐methyladenosine (m6A), as a eukaryotic mRNA modification catalyzed by methyltransferase METTL3, is involved in various processes of development or diseases via regulating RNA metabolism. However, the effect of METTL3‐mediated m6A modification in tooth development has remained elusive. Here we show that METTL3 is prevalently expressed in odontoblasts, dental pulp cells, dental follicle cells, and epithelial cells in Hertwig's epithelial root sheath during tooth root formation. Depletion of METTL3 in human dental pulp cells (hDPCs) impairs proliferation, migration, and odontogenic differentiation. Furthermore, conditional knockout of Mettl3 in Osterix‐expressing cells leads to short molar roots and thinner root dentin featured by decreased secretion of pre‐dentin matrix and formation of the odontoblast process. Mechanistically, loss of METTL3 cripples the translational efficiency of the key root‐forming regulator nuclear factor I‐C (NFIC). The odontogenic capacity of METTL3‐silenced hDPCs is partially rescued via overexpressing NFIC. Our findings suggest that m6A methyltransferase METTL3 is crucial for tooth root development, uncovering a novel epigenetic mechanism in tooth root formation. © 2020 American Society for Bone and Mineral Research (ASBMR).