Molecular cloning and expression analysis of carp (Cyprinus carpio) interleukin-1β, high affinity immunoglobulin E Fc receptor γ subunit and serum amyloid A

Abstract

Suppression subtractive hybridisation (SSH) is a powerful means to identify genes of cytokines and other genes that express small amount of mRNA. In this study, cDNA of normal fish (carp) head kidney cells (HKC) was subtracted from pooled cDNA of HKC and peritoneal cell (PC) obtained from fish which had been injected with sodium alginate (SA) and scleroglucan (SG) 3–48h earlier. This subtraction produced 248 clones of cDNA fragments. After sequencing some of the fragments of interest were used as probes, and yielded full-length cDNAs homologous to mammalian interleukin-1β (IL-1β), the γ subunit of high affinity Fc receptor for IgE (FcϵRIγ) and serum amyloid A (SAA); these were cloned and sequenced. Carp IL-1β shows 21·8–24·7% amino acid identities to mammalian mature IL-1β, and lacks a signal sequence, which is consistent with mammalian IL-1β. Carp FcϵRIγ, which was the first cloned non-mammalian Fc receptor subunit, shows 39·3–40·4% amino acid identities to mammalian FcϵRIγ, and contains the immunoreceptor tyrosin-based activation motif characteristic of the signal transduction subunit of antigen- and Fc-receptors. Carp SAA is most similar to mammalian acute phase responsive type SAA with 53·0–55·3% amino acid identities. Both SA-elicited and SG-elicited PC expressed higher amounts of IL-1β and SAA mRNA compared to saline-injected fish HKC and PC, indicating that these proteins are associated with inflammatory responses, similar to mammalian homologues. FcϵRIγ was constitutively expressed in leucocytes and not immunopotentiator-responsive, but this indicates that Fc receptor including FcϵRIγ subunit is likely functional in the carp immune system.