Essential Role of IL-4 and IL-4Rα Interaction in Adaptive Immunity of Zebrafish: Insight into the Origin of Th2-like Regulatory Mechanism in Ancient Vertebrates

Abstract The roles of IL-4 and IL-4Rα in Th2-mediated immunity have been well characterized in humans and other mammals. In contrast, few reports have been documented in ancient vertebrates. Several putative IL-4– and IL-4Rα–like molecules were identified recently from a few fish species, providing preliminary insight into the occurrence of Th2-type immunity in teleosts. However, functional determination still is required to address this hypothesis. To this end, these two molecules were characterized functionally in zebrafish (Danio rerio). Besides the identification of a full-length IL-4Rα molecule and an isoform lacking most of the cytoplasmic region as predicted previously, two novel alternatively spliced soluble variants with the extracellular domain only also were identified. Zebrafish IL-4Rα (DrIL-4Rα) shared overall conserved structural features of the IL-4Rα family. Immunofluorescence staining showed that DrIL-4Rα distributed on B cells. In vitro binding assays demonstrated that zebrafish IL-4 (DrIL-4) can bind specifically to DrIL-4Rα. In vivo administration of DrIL-4 significantly upregulated B cell proliferation and Ab production. These DrIL-4–elicited immune responses were downregulated by the administration of zebrafish soluble IL-4Rα or by DrIL-4Rα blockade using anti–DrIL-4Rα Abs. In addition, Th2-related cytokines or transcription factors were upregulated by DrIL-4. The DrIL-4–DrIL-4Rα interaction promoted CD40 expression on B cells and enhanced the CD154–CD40 costimulatory response, both of which are crucial for the initiation of Th2-type immunity. To our knowledge, this is the first report showing that a possible Th2-mediated regulatory mechanism may have appeared before the divergence of teleosts and mammals. These results add greater insight into the evolutionary history of adaptive immunity.