An important question in stem cell biology is how a cell decides to self-renew or differentiate. Drosophila neuroblasts divide asymmetrically to self-renew and generate differentiating progeny called GMCs. Here, we report that the Brain tumor (Brat) translation repressor is partitioned into GMCs via direct interaction with the Miranda scaffolding protein. In brat mutants, another Miranda cargo protein (Prospero) is not partitioned into GMCs, GMCs fail to downregulate neuroblast gene expression, and there is a massive increase in neuroblast numbers. Single neuroblast clones lacking Prospero have a similar phenotype. We conclude that Brat suppresses neuroblast stem cell self-renewal and promotes neuronal differentiation.

The Drosophila brain tumor (brat) gene encodes a member of the conserved NHL family of proteins, which appear to regulate differentiation and growth in a variety of organisms. One of the founding family members, Caenorhabditis elegans LIN-41, is thought to control posttranscriptional gene expression. However, the mechanism by which LIN-41, or any other NHL protein, acts has not been clear. Using a yeast "four-hybrid" interaction assay, we show that Brain Tumor is recruited to hunchback (hb) mRNA through interactions with Nanos and Pumilio, which bind to the RNA to repress its translation. Interaction with the Nanos/Pumilio/ RNA complex is mediated by the Brat NHL domain; single amino acid substitutions in this domain compromise quaternary complex assembly in vitro and hb regulation in vivo. Thus, recruitment of Brat is necessary for translational repression and the normal development of posterior embryonic pattern. In addition to regulating abdominal segmentation, previous genetic analysis has shown that Brat, Nanos, and Pumilio govern a variety of developmental processes. We examined the role of Brat in two of these processesregulation of maternal Cyclin B mRNA in the embryo and regulation of imaginal disc development. The results of these experiments suggest that NHL domain proteins are recruited to various mRNAs by combinatorial protein-protein interactions.

Posterior patterning in Drosophila embryos is governed by nanos (nos), which acts by repressing the translation of maternal transcripts of the hunchback (hb) gene. Sites in hb mRNA that mediate this repression, named nanos response elements (NREs), have been identified. However, we know of no evidence of a direct interaction between nos, or any other protein, and the NRE. Here, we show that two proteins present in embryonic extracts, neither one nos, bind specifically to the NRE in vitro. Furthermore, we show that binding in vitro correlates with NRE function in vivo. One of the NRE-binding factors is encoded by pumilio (pum), a gene that, like nos, is essential for abdominal segmentation. These and other observations suggest that pum acts by recognizing the NRE and then recruiting nos. Presumably, the resulting complex inhibits some component of the translation machinery.