Gasdermin family proteins are important effector proteins mediating pyroptosis and play an important role in innate immune response. GSDME can be cleaved by inflammatory Caspases at specific sites, releasing an active form of N-terminal fragment that binds to the plasma membrane to form pores and release cellular contents. Here, two GSDME genes, CcGSDME-like (CcGSDME-L) and CcGSDMEa, were cloned from common carp. The sequence similarity of the two genes were very high and more similar to DrGSDMEa of zebrafish in evolution. The expression levels of CcGSDME-L and CcGSDMEa can respond to the stimulation of Edwardsiella tarda. The results of cytotoxicity assay showed that CcGSDMEs were cleaved by the activation of canonical CcNLRP1 inflammasome, leading to obvious pyroptosis characteristics and increased cytotoxicity. In EPC cells, three CcCaspases responded to intracellular LPS stimulation and induced significantly cytotoxicity. In order to clarify the molecular mechanism of CcGSDME-induced pyroptosis, the N-terminal of CcGSDME-L (CcGSDME-L-NT) was expressed in 293T cells, which showed strong cytotoxicity and obvious pyroptosis characteristics. Fluorescence localization assay showed that the CcGSDME-L-NT was expressed on cell membrane, and CcGSDMEa-NT was located on the cell membrane or some organelle membranes. These findings can enrich the knowledge of CcNLRP1 inflammasome and GSDMEs mediated pyroptosis in common carp, and provide basic data for the prevention and treatment of fish infectious diseases.
Keywords: Common carp (Cyprinus carpio L.); GSDMEs; Inflammasome; Innate immunity; Pyroptosis.
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