Cloning and Characterization of Two Inhibitors of Apoptosis (IAP) from Eriocheir hepuensis and Their Expression Pattern in the Immune Organs under Azadirachtin Stress

Abstract

Inhibitors of apoptosis proteins (IAPs) maintain a balance between cell growth and cell death by suppressing caspase activity and modulating immunological responses. Here, two IAPs (EhIap1 and EhIap2) were identified from Eriocheir hepuensis using RT-PCR technology and characterized using bioinformatics software. Sequence analysis showed that the open reading frames of EhIap1 and EhIap2 were composed of 1,839 bp and 696 bp and encoded 612 amino acids and 231 amino acids, respectively. EhIap1 contained two BIR domains, one UBA domain, and one RING domain, and EhIap2 only contained two BIR domains. qRT-PCR showed that EhIap1 and EhIap2 were expressed in various tissues, and their expression levels were highest in the hepatopancreas. Expression levels of EhIAP1 and Eh1ap2 were more than 87.09 and 411 times higher in the hepatopancreas than in muscle, respectively. Under azadirachtin stress, the expression of EhIap1 and EhIap2 was increased in three immune tissues (hepatopancreas, gill, and heart). The expression of EhIap1 was 18.44, 17.75, and 6.16 times higher in hepatopancreas, heart, and gill tissues under azadirachtin stress compared with the control group, respectively. The expression of EhIap2 was 18.44, 15.48, and 3.91 times higher in hepatopancreas, gill, and heart tissue under azadirachtin stress compared with the control group, respectively. The expression of EhIap1 and EhIap2 was higher in surviving individuals than in dead individuals. Overall, EhIap1 and EhIap2 genes were first cloned, and our findings indicate that EhIAPs might be involved in E. hepuensis against azadirachtin through regulating apoptosis.