Exosomal miR-224 contributes to hemolymph microbiota homeostasis during bacterial infection in crustacean

Abstract

The modulation of hemolymph microbiota homeostasis is vital for the marine invertebrate innate immunity, while growing evidence shows that exosomes could serves as anti-bacterial immune factors, however, the relationship between exosomes and hemolymph microbiota homeostasis during pathogenic bacteria infection has not been addressed. Here, we determined that exosomes released from Vibrio parahaemolyticus-infected Scylla paramamosain (mud crabs) could reduce the mortality of the host during the infection by maintaining the homeostasis of hemolymph microbiota. We further confirmed that miR-224 was densely packaged in these exosomes and targeting to HSP70, which resulted in disruption of the HSP70-TRAF6 complex to release TRAF6 that allows it to interact with Ecsit. The interaction of TRAF6 with Ecsit regulates the production of mitochondrial ROS (mROS) and Anti-lipopolysaccharide factors (ALFs) expression in recipient hemocytes, which affects homeostasis of hemolymph microbiota in response to the pathogenic bacteria infection in mud crab. To the best of our knowledge, this is the first document that reports the role of exosome in the homeostasis of hemolymph microbiota during pathogen infection and a novel regulatory mechanism and crosstalk between exosomal miRNAs and innate immune response in crustaceans. Author summaryExosomes are small membrane vesicles of endocytic origin which are widely involved in the regulation of a variety of pathological processes in mammals. Yet, although the antibacterial function of exosomes has been discovered for many years, the relationship between exosomes and hemolymph microbiota homeostasis remains unknown. In the present study, we identified the miRNAs packaged by exosomes that were possibly involved in Vibrio parahaemolyticus infection by modulating hemolymph microbiota homeostasis in crustacean mud crab Scylla paramamosain. Moreover, it was found that miR-224 was densely packaged in exosomes after Vibrio parahaemolyticus challenge, resulting in the suppression of HSP70 and disruption of the HSP70-TRAF6 complex in recipient hemocytes, then the released TRAF6 was further interacted with Ecsit to regulate ROS and ALFs levels, which eventually affected hemolymph microbiota homeostasis to cope with pathogenic bacteria infection. Our finding is the first to reveal the relationship between exosomes and hemolymph microbiota homeostasis in animals, which shows a novel molecular mechanism of invertebrate resistance to pathogenic microbial infection.