Evaluation of siRNA-mediated knockdown of heat shock protein 16.2 in adult Acropora cervicornis

Acropora cervicornis is a critically endangered Caribbean coral species facing a precipitous decline due to disease and thermal bleaching. To combat the loss of ecologically important reef-building corals, novel molecular approaches can be used to understand mechanisms of resilience and ultimately provide a means to modulate coral performance. In this study, we tested a recently developed method for gene knockdown in adult corals that utilizes short interfering RNAs (siRNAs) and the RNA interference pathway. The Heat Shock Protein 16.2 (HSP16.2) gene in fragments of adult A. cervicornis was targeted for knockdown using a single siRNA construct. RT-qPCR analysis of HSP16.2 gene expression was performed between filtered seawater, no template control siRNA (NTC), and siRNA targeting HSP16.2 across four genotypes of A. cervicornis with known differences in thermal susceptibility (two heat-tolerant and two heat-susceptible genotypes). There was no significant difference in HSP16.2 gene expression due to the treatment of HSP16.2 siRNAs versus controls, but we did identify a single genotype-driven effect that corroborates evidence that A. cervicornis heat tolerance is driven largely by genotype. No difference in photophysiological performance or mortality was observed between experimental and control treatments. There were, however, some differences in mortality and HSP16.2 expression among genotypes. Our results reveal that siRNA-mediated knockdown was not successful for this species and gene and thus further optimization is needed prior to broader applicability of siRNA-mediated knockdown in corals. We provide an assessment of potential methodological improvements to support the development of future reverse-genetics studies in corals.