Transcriptional regulation of the basic helix‐loop‐helix factor AmeloD during tooth development

Abstract

The epithelial-mesenchymal interactions are essential for the initiation and regulation of the development of teeth. Following the initiation of tooth development, numerous growth factors are secreted by the dental epithelium and mesenchyme that play critical roles in cellular differentiation. During tooth morphogenesis, the dental epithelial stem cells differentiate into several cell types, including inner enamel epithelial cells, which then differentiate into enamel matrix-secreting ameloblasts. Recently, we reported that the novel basic-helix-loop-helix transcription factor, AmeloD, is actively engaged in the development of teeth as a regulator of dental epithelial cell motility. However, the gene regulation mechanism of AmeloD is still unknown. In this study, we aimed to uncover the mechanisms regulating AmeloD expression during tooth development. By screening growth factors that are important in the early stages of tooth formation, we found that TGF-β1 induced AmeloD expression and ameloblast differentiation in the dental epithelial cell line, SF2. TGF-β1 phosphorylated ERK1/2 and Smad2/3 to induce AmeloD expression, whereas treatment with the MEK inhibitor, U0126, inhibited AmeloD induction.Promoter analysis of AmeloD revealed that the proximal promoter of AmeloD showed high activity in dental epithelial cell lines, which was enhanced following TGF-β1 stimulation. These results suggested that TGF-β1 activates AmeloD transcription via ERK1/2 phosphorylation. Our findings provide new insights into the mechanisms that govern tooth development.

Deficiency of G protein‐coupled receptor Gpr111/Adgrf2 causes enamel hypomineralization in mice by alteration of the expression of kallikrein‐related peptidase 4 (Klk4) during pH cycling process

“…cDNA was synthesized from 1 μg of the total RNA using the SuperScript™ VILO™ Master Mix (Invitrogen). Real‐time PCR was performed using SYBR™ Select Master Mix (Invitrogen) on a StepOnePlus™ Real‐time PCR system (Thermo Fisher Scientific), as previously described 24 . Briefly, 100 ng of cDNA and 200 nM of each primer were used for 20 μL of PCR reaction.…”

“…Real-time PCR was performed using SYBR™ Select Master Mix (Invitrogen) on a StepOnePlus™ Realtime PCR system (Thermo Fisher Scientific), as previously described. 24 Briefly, 100 ng of cDNA and 200 nM of each primer were used for 20 μL of PCR reaction. The results are representative of at least three independent experiments with triplicates per each experiment.…”

S100a6 knockdown promotes the differentiation of dental epithelial cells toward the epidermal lineage instead of the odontogenic lineage

“…Ameloblasts express extracellular factors such as Ambn, amelogenin (Amelx), AmeloD, von Willebrand factor D, and epidermal growth factor domains during the secretory phase. [4][5][6][7] Calcification is performed using the enamel matrix as a scaffold, 4,5 which is later degraded by proteases such as matrix metalloproteinase-20 and kallikrein-related peptidase 4 (Klk4) to enhance calcification. Although extensive studies have been performed on the differentiation of the dental epithelium into mature ameloblasts, 8,9 the calcification process remains poorly understood.…”

The tooth‐specific basic helix‐loop‐helix factor AmeloD promotes differentiation of ameloblasts

“…During tooth development, AmeloD is expressed in IEE cells until ameloblast differentiation occurs (He et al, 2018). The transcription of AmeloD is regulated by the transforming growth factor‐signaling pathway via extracellular signal‐regulated kinase 1/2 phosphorylation (Al Thamin et al, 2021). AmeloD binds to the E‐box cis‐element of E‐cadherin and downregulates its transcriptional activity; mice lacking AmeloD develop small teeth due to reduced dental epithelial cell motility (Chiba et al, 2019).…”

“…During tooth development, AmeloD is expressed in IEE cells until ameloblast differentiation occurs (He et al, 2018). The transcription of AmeloD is regulated by the transforming growth factor-signaling pathway via extracellular signalregulated kinase 1/2 phosphorylation (Al Thamin et al, 2021).…”