Functional activity and developmental regulation of DdRBP1, a RNA binding protein in Dictyostelium discoideum

In an attempt to find potential components of natural antisense mechanisms in Dictyostelium, we investigated RNA binding protein (RBD) genes of the RNP-CS family. RBD proteins can enhance hybridization of complementary RNAs and may thus mediate the interaction of sense and antisense RNA. Using the conserved RNP1 and RNP2 motifs as primers, we cloned 4 PCR fragments containing ORFs and additional homologies to known members of the RNP-CS family. We cloned a full length cDNA for one protein (DdRBP1) that showed similarities to hnRNP A1. Recombinant protein synthesized in E. coli displayed binding to single stranded RNA and a weak annealing activity for partially complementary RNAs in vitro. Deletion of the RNP1 motif reduced RNA binding considerably but not completely. DdRBP1 is thus one of the few members of the RNP-CS family for which binding and annealing activities have been experimentally demonstrated. Polyclonal antisera directed against recombinant DdRBP1 detected a protein of approx. 40 kDa. In whole cell extracts, this protein was present in equal amounts throughout the developmental cycle of Dictyostelium while differential accumulation was observed in nuclei during early and late development.