Molecular characteristics of AMPK and its role in regulating the phagocytosis of oyster hemocytes

Abstract

Phagocytosis is one of the fundamental cellular immune defense parameter that helps in the elimination of the invading pathogens in both vertebrates and invertebrates, which require plenty of energy for functioning. In the present study, we identified the critical energy regulator AMP-activated protein kinase (AMPK) in Crassostrea hongkongensis which is composed of three subunits, named ChAMPK-α, ChAMPK-β, and ChAMPK-γ, and then analyzed the function of AMPK in regulating hemocyte phagocytosis. All the three ChAMPK subunits mRNA were detected to be expressed at various embryological stages, and also constitutively expressed in multiple tissues with high expression in gill and mantle. The phylogenetic tree showed that the three subunits of AMPK were correspondingly clustered with its orthologue branches. Furthermore Western Blot analysis revealed that the AMPK pharmacological inhibitors Compound C could effectively down-regulate the Thr172 phosphorylation level of AMPK-α, and the hemocyte phagocytosis was inhibited by Compound C (CC), which indicate its existence in the oyster. Our results showed that treatment of AMPK inhibitors significantly attenuated the capacity of hemocytes phagocytosis. Moreover, Compound C could also change the organization of actin cytoskeleton in the oyster hemocytes, demonstrating the crucial role of AMPK signaling in control of phagocytosis.