The Neural RNA-Binding Protein Musashi1 Translationally Regulates Mammalian numb Gene Expression by Interacting with Its mRNA

Posttranscriptional regulation plays essential roles in the wide variety of events that occur during animal development, including the localization of maternal-effect gene products within oocytes, cell fate determination, and cell polarity formation by controlling alternative splicing, mRNA stability, RNA transport, and/or the translation of existing mRNAs (11, 57, 66). Among developing tissues, the nervous system in particular uses a variety of posttranscriptional means to regulate the vast cellular diversity and synaptic plasticity that is generated. Neural RNA-binding proteins are likely to play essential roles in mediating these processes (44). Two classes of neural RNA-binding proteins with ribonucleoprotein-type RNA recognition motifs (RRMs), the Elav and Musashi subfamilies, have been proposed (44). The Elav subfamily includes the mammalian Elav homologue, Hu proteins, whose members are expressed in postmitotic neurons and have been proposed to be required for the survival or terminal differentiation of these cells (1, 2, 34, 45, 64, 69). Intensive study in this area revealed that Hu proteins regulate the gene expression involved in neuronal differentiation by controlling RNA stabilization or translation (2, 13, 35, 48, 64). On the other hand, the Musashi subfamily proteins are expressed in neural precursor cells rather than postmitotic neurons (15, 28, 44, 49, 52). Musashi1 (Msi1) was isolated as a mammalian homologue of Drosophila Musashi (d-Msi), which is required for the asymmetric cell division of sensory neural precursor cells (38). In Drosophila, genes that are responsible for the proper asymmetric cell division of sensory organ precursor cells or central nervous system neuroblasts have been identified and extensively investigated. Interestingly, many of these genes have been shown to be involved in the regulation of Notch signaling, including numb, tramtrack, Notch, and Delta (reviewed by Jan and Jan [23]). Recently, our laboratory showed that d-Msi represses the expression of tramtrack, which encodes a transcriptional repressor, at the level of translation (21, 43). We also identified the mammalian homologue of Drosophila Musashi, Musashi1, which is highly enriched in neural progenitor cells in the developing mouse embryonic central nervous system (28, 52, 53). Msi1 expression is gradually down-regulated during the course of neural differentiation. The Msi1 protein consists of 362 amino acid (aa) residues, and it contains two conserved RRMs in its N-terminal half and a putative nuclear export signal in its C-terminal half, which is consistent with its observed localization of the cytoplasm in embryonic neural progenitor cells (28, 52). To help elucidate the role of Msi1 protein in neural progenitor cells, we sought to determine (i) the RNA sequences that bind to Msi1, (ii) an in vivo target RNA of Msi1, and (iii) the mechanism of action of Msi1 on the expression of its downstream target genes. To this end, in the present study we identified the RNA sequence for Msi1 and demonstrated putative translational repression of a likely in vivo target gene mammalian numb (m-numb).