In Vivo Analysis of Ifn-g1 and Ifn-g2 Signaling in Zebrafish

The zebrafish genome contains a large number of genes encoding potential cytokine receptor genes as judged by homology to mam-malian receptors. The sequences are too divergent to allow unambiguous assignments of all receptors to specific cytokines, and only a few have been assigned functions by functional studies. Among receptors for class II helical cytokines—i.e., IFNs that include virus-induced Ifns (Ifn-w) and type II Ifns (Ifn-g), together with Il-10 and its related cytokines (Il-20, Il-22, and Il-26)—only the Ifn-w–specific complexes have been functionally identified, whereas the receptors for the two Ifn-g (Ifn-g1 and Ifn-g2) are unknown. In this work, we identify conditions in which Ifn-g1 and Ifn-g2 (also called IFNG or IFN-g and IFN-gammarel) are induced in fish larvae and adults. We use morpholino-mediated loss-of-function analysis to screen candidate receptors and identify the components of their receptor complexes. We find that Ifn-g1 and Ifn-g2 bind to different receptor complexes. The receptor complex for Ifn-g2 includes cytokine receptor family B (Crfb)6 together with Crfb13 and Crfb17, whereas the receptor complex for Ifn-g1 does not include Crfb6 or Crfb13 but includes Crfb17. We also show that of the two Jak2 paralogues present in the zebrafish Jak2a but not Jak2b is involved in the intracellular transmission of the Ifn-g signal. These results shed new light on the evolution of the Ifn-g signaling in fish and tetrapods and contribute toward an integrated view of the innate immune regulation in vertebrates. I nterferons are members of the large family of helical cyto-kines. These cytokines are structured around a four a helix bundle and are divided into class I cytokines, which include hormones of the prolactin and growth hormone family together with erythropoietin, leptin, and the majority of ILs (1), and class II cytokines. Class II cytokines are represented by the IFNs together with IL-10 and its relatives (IL-20, IL-22, IL-24, and IL-26). In mammals, the IFN-subclass is divided into three types: type I (IFN-a/b/v/ε/k) and type III (IFN-l) are the main players in innate immunity against viral infections. Type II, with its only representative IFN-g, has clear antiviral activity but is also an immune regulator required for resistance to a wide range of bacterial and protozoan pathogens (e.g., Salmonella, leishmania, and trypano-somes) in mice. In man, it appears to be instrumental in resistance to mycobacterial diseases (2, 3) but also is involved in Ag presentation (4), cancer immunoediting (5), leukocyte–endothelium interactions (2), and osteoclastogenesis suppression (6, …