Molecular and functional characterization of erythropoietin receptor of the goldfish (Carassius auratus L.)

Abstract

Erythropoietin receptor (EPOR) is a member of the class I cytokine receptor superfamily and signaling through this receptor is important for the proliferation, differentiation and survival of erythrocyte progenitor cells. This study reports on the molecular and functional characterization of goldfish EPOR. The identified goldfish EPOR sequence possesses the conserved EPOR ligand binding domain, the fibronectin domain, the class I cytokine receptor superfamily motif (WSXWS) as well as several intracellular signaling motifs characteristic of other vertebrate EPORs. The expression of epor mRNA in goldfish tissues, cell populations and cells treated with recombinant goldfish EPO (rgEPO) were evaluated by quantitative PCR revealing that goldfish epor mRNA is transcribed in both erythropoietic tissues (blood, kidney and spleen) and non-hematopoietic tissues (brain, heart and gill), as well as in immature erythrocytes. Recombinant goldfish EPOR (rgEPOR), consisting of its extracellular domain, dose-dependently inhibited proliferation of progenitor cells induced by rgEPO. In vitro binding studies indicated that rgEPO exists as monomer, dimer and/or trimmer and that rgEPOR exists as monomer and/or homodimer, and when incubated together, formed a ligand–receptor complex. Our results demonstrate that goldfish EPO/EPOR signaling has been highly conserved throughout vertebrate evolution as a required mechanism for erythrocyte development.