Molecular characteristics and the roles of CaASC and its restriction to Aeromonas hydrophila in Carassius auratus.

Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), as a critical adaptor molecule in inflammasome complexes, plays an important role in mediating inflammation reaction. In this study, the complete cDNA of ASC gene with 891 bp was cloned in Qihe crucian carp Carassius auratus (named as CaASC), which was composed of a 5'-UTR of 36 bp, a 3'-UTR of 252 bp, and an ORF of 603 bp encoded 200 amino acids with a predicted isoelectric point of 5.61 and a molecular mass of 22.0 kDa. Multiple sequence alignment and motif analysis revealed that CaASC contained a conserved N-terminal Pyrin domain (PYD) and a C-terminal Caspase recruitment domain (CARD). CaASC mRNA and protein expressions were detected in selected tissues, with the highest mRNA level in the spleen. Meanwhile, CaASC gene expressions were clearly altered in intestine, gill, skin, spleen, liver and head kidney of fish challenged by Aeromonas hydrophila, LPS, and polyI:C, respectively. The recombined proteins of CaASC with fluorescent tag were over-expressed in transfected 293T cells, and the green specks were observed obviously and located in the cytoplasm. Furthermore, knockdown of CaASC reduced the expression of IL-1ß and promoted the bacterial colonization in fish tissues, while overexpression of CaASC increased the expression of IL-1ß and hampered the bacterial colonization in these tissues. Taken together, these results identified the molecular characteristics of CaASC in C. auratus, and revealed its role in regulating IL-1ß expression and restricting bacterial infection in vivo.