Drosophila YBX1 homolog YPS promotes ovarian germline stem cell development by preferentially recognizing 5-methylcytosine RNAs

5-Methylcytosine (m5C) is one of the most abundant RNA modifications, but its function in adult stem cell development remains poorly defined. This study shows that Ypsilon schachtel (YPS) promotes germ line stem cell (GSC) maintenance, proliferation, and differentiation in the adult Drosophila ovary by preferentially binding to m5C-containing RNAs. Highly conserved cold-shock domains (CSDs) of YPS and its human homolog Y box binding protein 1 (YBX1) exhibit preferential binding to m5C-containing RNAs through hydrophobic interactions. Human YBX1 can functionally replace YPS to promote GSC development in the Drosophila ovary, and overexpressing RNA-binding–defective YBX1 and YPS mutant proteins disrupts normal GSC development. Thus, this study provides insight into the importance of m5C RNA modification in adult stem cell development. Overall design: Pull down m5C RNA with specific 5-methylcytosine (5-mC) monoclonal antibody, pull down RNA/Protein complexes via Flag tagged Yps overexpressed proteins.