von Willebrand factor D and EGF domains regulate ameloblast differentiation and enamel formation

Abstract

Cell‐ and tissue‐specific extracellular matrix (ECM) composition plays an important role in organ development, including teeth, by regulating cell behaviors, such as cell proliferation and differentiation. Here, we demonstrate for the first time that von Willebrand factor D and epidermal growth factor (EGF) domains (Vwde), a previously uncharacterized ECM protein, is specifically expressed in teeth and regulates cell proliferation and differentiation in inner enamel epithelial cells (IEEs) and enamel formation. We identified the Vwde as a novel ECM protein through bioinformatics using the NCBI expressed sequence tag database for mice. Vwde complementary DNA encodes 1773 amino acids containing a signal peptide, a von Willebrand factor type D domain, and tandem calcium‐binding EGF‐like domains. Real‐time polymerase chain reaction demonstrated that Vwde is highly expressed in tooth tissue but not in other tissues including the brain, lung, heart, liver, kidney, and bone. In situ hybridization revealed that the IEEs expressed Vwde messenger RNA in developing teeth. Immunostaining showed that VWDE was localized at the proximal and the distal ends of the pericellular regions of the IEEs. Vwde was induced during the differentiation of mouse dental epithelium‐derived M3H1 cells. Vwde‐transfected M3H1 cells secreted VWDE protein into the culture medium and inhibited cell proliferation, whereas ameloblastic differentiation was promoted. Furthermore, Vwde increased the phosphorylation of extracellular signal‐regulated kinase 1/2 and protein kinase B and strongly induced the expression of the intercellular junction protein, N‐cadherin (Ncad). Interestingly, the suppression of endogenous Vwde inhibited the expression of Ncad. Finally, we created Vwde‐knockout mice using the CRISPR‐Cas9 system. Vwde‐null mice showed low mineral density, rough surface, and cracks in the enamel, indicating the enamel hypoplasia phenotype. Our findings suggest that Vwde assembling the matrix underneath the IEEs is essential for Ncad expression and enamel formation.

Chromosome-level, nanopore-only genome and allele-specific DNA methylation of Pallas's cat, Otocolobus manul

“…Genes overlapping DMRs included previously described imprinted loci such as (A) potassium voltage-gated channel subfamily Q member 1 (KCNQ1) and (B) the guanine nucleotide-binding protein G(S) subunit alpha (GNAS) complex locus (aliases LOC101098453, LOC102900772). DMRs also overlapped genes not previously reported as imprinted, including (C) von Willebrand factor D and EGF domains (VWDE), a protein with predicted involvement in anatomical development ( 96 ). (D) The most notable locus was a 33.2 kb region containing 12 DMRs all hypomethylated on pseudohaplotype 2.…”

Comparative genomics revealed drastic gene difference in two small Chinese perches, Siniperca undulata and S. obscura

“…Furthermore, many expanding gene families in S. undulata include Tenm1 and Vwdel genes. The Tenm1 gene can promote the generation of synapses among neural cells and connect neurons ( Li et al 2018 ), while the Vwdel gene regulates proliferation and differentiation of enamel epithelial cells in dental tissue ( Iwata et al 2022 ), and its expression is associated with the regeneration of lambs and fins ( Leigh et al 2020 ). Additionally, the rnf183 gene, which takes part in the regulation of the cell cycle and the promotion of terminal differentiation in the cell cycle ( Lu et al 2020b ), and the ltn1 gene, which plays an important role in the growth of the embryo ( Yang et al 2021 ), both positively selected in S. undulata .…”

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.…”

“…During tooth development, the dental epithelium differentiates into ameloblasts, which proceed through secretory, calcification, and maturation stages to form enamel. Ameloblasts express extracellular factors such as Ambn , amelogenin ( Amelx ), AmeloD, von Willebrand factor D, and epidermal growth factor domains during the secretory phase 4–7 . 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.…”