RNAi Silencing of the Biomineralization Gene Perlucin Impairs Oyster Ability to Cope with Ocean Acidification

Abstract

Calcifying marine organisms, including the eastern oyster (Crassostrea virginica), are vulnerable to ocean acidification (OA) because it is more difficult to precipitate calcium carbonate (CaCO3). Previous investigations of the molecular mechanisms associated with resilience to OA in C. virginica demonstrated significant differences in single nucleotide polymorphism and gene expression profiles among oysters reared under ambient and OA conditions. Converged evidence generated by both of these approaches highlighted the role of genes related to biomineralization, including perlucins. Here, gene silencing via RNA interference (RNAi) was used to evaluate the protective role of a perlucin gene under OA stress. Larvae were exposed to short dicer-substrate small interfering RNA (DsiRNA-perlucin) to silence the target gene or to one of two control treatments (control DsiRNA or seawater) before cultivation under OA (pH ~7.3) or ambient (pH ~8.2) conditions. Two transfection experiments were performed in parallel, one during fertilization and one during early larval development (6 h post-fertilization), before larval viability, size, development, and shell mineralization were monitored. Silenced oysters under acidification stress were the smallest, had shell abnormalities, and had significantly reduced shell mineralization, thereby suggesting that perlucin significantly helps larvae mitigate the effects of OA.