The complement system is important in the innate immune response. C1q-domain-containing proteins have multiple functions and occur extensively in invertebrates and vertebrates. In this study, PoC1ql3 encoding a C1q-domain-containing protein in the Japanese flounder was identified. The 266-amino-acid polypeptide encoded, PoC1ql3, shares high sequence and structural similarity with orthologues in other fish and mammals. PoC1ql3 is abundantly expressed in the brain, but less in the blood, gills, and liver. Transcripts of PoC1ql3 were down-regulated in the spleen and liver 6–24 h after bacterial infection, but were significantly up-regulated after 48 h. Full-length PoC1ql3 (C1ql3-full) and its gC1q domain (C1ql3-part) were both exerted anti-Edwardsiella tarda activity. C1ql3-part bound to lipopolysaccharide and peptidoglycan, and exerted antibacterial effects against E. tarda in vivo, suggesting that C1ql3 functions as a pathogen-recognition receptor. Therefore, PoC1ql3 functions in the innate immune system, which would facilitate the investigation of the immune system in Japanese flounder.