Abstract
AbstractKit ligand (Kitlg) is pleiotropic cytokine with a prominent role in vertebrate erythropoiesis. Although the role of Kitlg in this process has not yet been reported inDanio rerio(zebrafish), in the present study, we show that its function is evolutionary conserved. Zebrafish possess two copies of Kitlg genes (Kitlga and Kitlgb) due to whole genome duplication. To determine the role of each ligand in zebrafish, we performed a series ofex vivoandin vivogain- and loss-of-function experiments. First, we tested the biological activity of recombinant Kitlg proteins in suspension culture from zebrafish whole kidney marrow and we demonstrate that Kitlga is necessary for expansion of erythroid progenitorsex vivo. To further address the role ofkitlgaandkitlgbin hematopoietic developmentin vivo, we performed gain-of-function experiments in zebrafish embryos, showing that both ligands cooperate with erythropoietin (Epo) to promote erythroid cell expansion. Finally, using thekitamutant (kitab5/b5orsparse), we show that Kita receptor is crucial for Kitlga/b cooperation with Epo in erythroid cells. In summary, using optimized suspension culture conditions with recombinant cytokines (Epo, Kitlga), we are reporting for the first timeex vivosuspension cultures of zebrafish hematopoietic progenitor cells, which can serve as an indispensable tool to study normal and aberrant hematopoiesis in zebrafish. Furthermore, we conclude that although partial functional diversification of Kit ligands has been described in other processes, in erythroid development, both paralogs play a similar role and their function is evolutionary conserved.Key points:Kit signaling contributes to erythroid cell development and is conserved from fish to manEx vivo expansion and self-renewal of zebrafish erythroid progenitors requires addition of recombinant Kitlga
Publisher
Cold Spring Harbor Laboratory