Otopetrins comprise a family of proton-selective channels that are critically important for the mineralization of otoliths and statoconia in vertebrates but whose underlying cellular mechanisms remain largely unknown. Here, we demonstrate that otopetrins are critically involved in the calcification process by providing an exit route for protons liberated by the formation of CaCO₃. Using the sea urchin larva, we examined the otopetrin ortholog otop2l, which is exclusively expressed in the calcifying primary mesenchymal cells (PMCs) that generate the calcitic larval skeleton. otop2l expression is stimulated during skeletogenesis, and knockdown of otop2l impairs spicule formation. Intracellular pH measurements demonstrated Zn²⁺-sensitive H⁺ fluxes in PMCs that regulate intracellular pH in a Na⁺/HCO₃⁻-independent manner, while Otop2l knockdown reduced membrane proton permeability. Furthermore, Otop2l displays unique features, including strong activation by high extracellular pH (>8.0) and check-valve–like outwardly rectifying H⁺ flux properties, making it into a cellular proton extrusion machine adapted to oceanic living conditions. Our results provide evidence that otopetrin family proton channels are a central component of the cellular pH regulatory machinery in biomineralizing cells. Their ubiquitous occurrence in calcifying systems across the animal kingdom suggest a conserved physiological function by mediating pH at the site of mineralization. This important role of otopetrin family proton channels has strong implications for our view on the cellular mechanisms of biomineralization and their response to changes in oceanic pH.

Otopetrins 包含一个质子选择性通道家族，这些通道对于脊椎动物的耳石和静耳石的矿化至关重要，但其潜在的细胞机制仍然很大程度上未知。在这里，我们证明了 otopetrins 通过为 CaCO ₃的形成释放的质子提供出口途径，在钙化过程中发挥了关键作用。使用海胆幼虫，我们检查了 otopetrin ortholog otop2l，它仅在生成钙质幼虫骨架的钙化初级间充质细胞 (PMC) 中表达。otop2l表达在骨骼形成过程中受到刺激，并且otop2l的敲低会损害针状体的形成。细胞内 pH 值测量表明 Zn^{PMC 中的2+} -敏感 H ⁺通量以 Na ⁺ /HCO ₃ ^-非依赖性方式调节细胞内 pH 值，而 Otop2l 敲低降低了膜质子通透性。此外，Otop2l 表现出独特的特征，包括通过高细胞外 pH 值 (>8.0) 的强烈激活和单向阀状向外整流 H ⁺通量特性，使其成为适应海洋生活条件的细胞质子挤压机。我们的结果提供了证据，证明 otopetrin 家族质子通道是生物矿化细胞中细胞 pH 调节机制的核心组成部分。它们在动物界的钙化系统中普遍存在，表明通过调节矿化部位的 pH 值具有保守的生理功能。otopetrin 家族质子通道的这一重要作用对我们对生物矿化的细胞机制及其对海洋 pH 值变化的反应的看法具有重要意义。