Ocean acidification disrupts the biomineralization process in the oyster Crassostrea virginica via intracellular calcium signaling dysregulation

Abstract Anthropogenically increased atmospheric carbon dioxide (pCO₂) leads to ocean acidification, disrupting calcification in marine calcifiers by reducing the saturation state of calcium carbonate. Calcium is not only a crucial component in the shell and skeleton structure but also serves as an essential second messenger for regulating biomineralization across many species. Ocean acidification is well-studied as causing shell dissolution in a diversity of bivalve species by disordering calcium deposition. However, it remains unclear whether the calcium-mediated signaling pathway regulating biomineralization is also affected. This study assessed eastern oyster (Crassostrea virginica) to determine how calcium signaling responds to elevated pCO₂ and influences shell formation. Under elevated pCO₂, increased intracellular calcium concentration was found in primary epithelial cell cultures from oyster mantle. Meanwhile, we observed upregulation of calmodulin, a primary sensor of intracellular calcium, while its downstream effector, calcineurin, was downregulated. In addition, four conserved shell matrix proteins (SMPs), representing shell construction conditions, were significantly upregulated in the CO₂-exposed mantle cells. In vivo, larval C. virginica exhibited developmental stage-dependent alterations in calcium signaling and SMPs disarrangement stimulated by pCO₂. We hypothesize that dysregulation of calcium signaling disrupts the expressions of SMPs and causes oyster shell deformation. Pharmaceutical blockage of the calcium-calmodulin binding induced abnormal expression of related genes and shell matrix changes consistent with those caused by elevated pCO₂, both in vivo and in vitro. Importantly, calcineurin restored SMPs expression in CO₂-treated mantle cells. These findings suggest that shell deformities under ocean acidification are related to disruption of the calcium-calmodulin signaling pathway, inhibiting calcineurin activity and affecting SMPs production.