LPA₆-RhoA signals regulate junctional complexes for polarity and morphology establishment of maturation stage ameloblasts

Objectives: Lysophosphatidic acid (LPA) is a potent bioactive phospholipid that exerts various functions upon binding to six known G protein-coupled receptors (LPA_1-6); however; its role in a tooth remains unclear. This study aimed to explore the impact of the LPA/LPA receptor 6 (LPA₆)/RhoA signaling axis on maturation stage ameloblasts (M-ABs), which are responsible for enamel mineralization.

Methods: The expression of LPA₆ and LPA-producing synthetic enzymes during ameloblast differentiation was explored through immunobiological analysis of mouse incisors and molars. To elucidate the role of LPA₆ in ameloblasts, incisors of LPA₆ KO mice were analyzed. In vitro experiments using ameloblast cell lines were performed to validate the function of LPA-LPA₆-RhoA signaling in ameloblasts.

Results: LPA₆ and LPA-producing enzymes were strongly expressed in M-ABs. In LPA₆ knockout mice, M-ABs exhibited abnormal morphology with the loss of cell polarity, and an abnormal enamel epithelium containing cyst-like structures was formed. Moreover, the expression of E-cadherin and zonula occludens-1 (ZO-1) significantly decreased in M-ABs. In vitro experiments demonstrated that LPA upregulated the expression of E-cadherin, ZO-1, and filamentous actin (F-actin) at the cellular membrane, whereas LPA₆ knockdown decreased their expression and changed cell morphology. Furthermore, we showed that RhoA signaling mediates LPA-LPA₆-induced junctional complexes.

Conclusions: This study demonstrated that LPA-LPA₆-RhoA signaling is essential for establishing proper cell morphology and polarity, via cell-cell junction and actin cytoskeleton expression and stability, of M-ABs. These results highlight the biological significance of bioactive lipids in a tooth, providing a novel molecular regulatory mechanism of ameloblasts.