Widespread regulation of the maternal transcriptome by Nanos in Drosophila.

The translational repressor Nanos (Nos) regulates a single target, maternal hunchback (hb) mRNA, to govern abdominal segmentation in the early Drosophila embryo. Nos is recruited to sites in the 3' UTR of hb mRNA in collaboration with the sequence-specific RNA-binding protein Pumilio (Pum); on its own, Nos has no binding specificity. Nos is expressed at other stages of development, but very few mRNA targets that might mediate its action at these stages have been described.

Significant roles in RNA-binding for the amino-terminal domains of Drosophila Pumilio and Nanos.

The Drosophila Pumilio (Pum) and Nanos (Nos) RNA-binding proteins govern abdominal segmentation in the early embryo, as well as a variety of other events during development. They bind together to a compound Nanos Response Element (NRE) present in thousands of maternal mRNAs in the ovary and embryo, including ( ) mRNA, thereby regulating poly-adenylation, translation, and stability. Many studies support a model in which mRNA recognition and effector recruitment are achieved by distinct regions of each protein.

Widespread regulation of the maternal transcriptome by Nanos in Drosophila.

The translational repressor Nanos (Nos) regulates a single target, maternal hunchback (hb) mRNA, to govern abdominal segmentation in the early Drosophila embryo. Nos is recruited specifically to sites in the 3'-UTR of hb mRNA in collaboration with the sequence-specific RNA-binding protein Pumilio (Pum); on its own, Nos has no binding specificity. Nos is expressed at other stages of development, but very few mRNA targets that might mediate its action at these stages have been described.

No significant regulation of bicoid mRNA by Pumilio or Nanos in the early Drosophila embryo.

Drosophila Pumilio (Pum) is a founding member of the conserved Puf domain class of RNA-binding translational regulators. Pum binds with high specificity, contacting eight nucleotides, one with each of the repeats in its RNA-binding domain. In general, Pum is thought to block translation in collaboration with Nanos (Nos), which exhibits no binding specificity in isolation but is recruited jointly to regulatory sequences containing a Pum binding site in the 3'-UTRs of target mRNAs.

E(nos)/CG4699 required for nanos function in the female germ line of Drosophila.

The translational repressor Nanos is required in the germ line stem cells of the Drosophila ovary to maintain their capacity for self-renewal. Following division of the stem cells, Nanos is inhibited in the daughters that differentiate into cysts and ultimately become mature oocytes. The control of Nanos activity is thus an important aspect of the switch from self-renewal to differentiation.

Co-occupancy of two Pumilio molecules on a single hunchback NRE.

Pumilio controls a number of processes in eukaryotes, including the translational repression of hunchback (hb) mRNA in early Drosophila embryos. The Pumilio Puf domain binds to a pair of 32 nucleotide (nt) Nanos response elements (NRE1 and NRE2) within the 3' untranslated region of hb mRNA. Despite the elucidation of structures of human Pumilio Puf domain in complex with hb RNA elements, the nature of hb mRNA recognition remains unclear.

A splicer that represses (translation).

Regulated translation and subcellular localization of maternal mRNAs underlies establishment of the antero-posterior axis in the Drosophila oocyte. In this issue of Genes & Development, Besse et al. (pp.

Structures of human Pumilio with noncognate RNAs reveal molecular mechanisms for binding promiscuity.

Pumilio is a founder member of the evolutionarily conserved Puf family of RNA-binding proteins that control a number of physiological processes in eukaryotes. A structure of human Pumilio (hPum) Puf domain bound to a Drosophila regulatory sequence showed that each Puf repeat recognizes a single nucleotide. Puf domains in general bind promiscuously to a large set of degenerate sequences, but the structural basis for this promiscuity has been unclear.

The molecular chaperone Hsp90 is required for mRNA localization in Drosophila melanogaster embryos.

Localization of maternal nanos mRNA to the posterior pole is essential for development of both the abdominal segments and primordial germ cells in the Drosophila embryo. Unlike maternal mRNAs such as bicoid and oskar that are localized by directed transport along microtubules, nanos is thought to be trapped as it swirls past the posterior pole during cytoplasmic streaming. Anchoring of nanos depends on integrity of the actin cytoskeleton and the pole plasm; other factors involved specifically in its localization have not been described to date.

Translational control of maternal Cyclin B mRNA by Nanos in the Drosophila germline.

In the Drosophila embryo, Nanos and Pumilio collaborate to repress the translation of hunchback mRNA in the somatic cytoplasm. Both proteins are also required for repression of maternal Cyclin B mRNA in the germline; it has not been clear whether they act directly on Cyclin B mRNA, and if so, whether regulation in the presumptive somatic and germline cytoplasm proceeds by similar or fundamentally different mechanisms. In this report, we show that Pumilio and Nanos bind to an element in the 3' UTR to repress Cyclin B mRNA.

mRNA regulation by Puf domain proteins.

Puf domain proteins bind specific sequences in mRNAs to regulate their translation or stability, or both. Neither the mechanism of their action nor the identities of targeted mRNAs have been well defined. Recent work suggests that Puf proteins generally act by recruiting Pop2, a deadenylation enzyme that is part of a large complex.

Brat is a Miranda cargo protein that promotes neuronal differentiation and inhibits neuroblast self-renewal.

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.

Solution structure of the Vts1 SAM domain in the presence of RNA.

The yeast Vts1 SAM (sterile alpha motif) domain is a member of a new class of SAM domains that specifically bind RNA. To elucidate the structural basis for RNA binding, the solution structure of the Vts1 SAM domain, in the presence of a specific target RNA, has been solved by multidimensional heteronuclear NMR spectroscopy. The Vts1 SAM domain retains the "core" five-helix-bundle architecture of traditional SAM domains, but has additional short helices at N and C termini, comprising a small substructure that caps the core helices.

The translational repressor Pumilio regulates presynaptic morphology and controls postsynaptic accumulation of translation factor eIF-4E.

Translational repression by Drosophila Pumilio (Pum) protein controls posterior patterning during embryonic development. Here, we show that Pum is an important mediator of synaptic growth and plasticity at the neuromuscular junction (NMJ). Pum is localized to the postsynaptic side of the NMJ in third instar larvae and is also expressed in larval neurons.

Degradation of origin recognition complex large subunit by the anaphase-promoting complex in Drosophila.

The initiation of DNA synthesis is thought to occur at sites bound by a heteromeric origin recognition complex (ORC). Previously, we have shown that in Drosophila, the level of the large subunit, ORC1, is modulated during cell cycle progression and that changes in ORC1 concentration alter origin utilization during development. Here, we investigate the mechanisms underlying cell cycle-dependent degradation of ORC1.

Model of the brain tumor-Pumilio translation repressor complex.

The Brain Tumor (Brat) protein is recruited to the 3' untranslated region (UTR) of hunchback mRNA to regulate its translation. Recruitment is mediated by interactions between the Pumilio RNA-binding Puf repeats and the NHL domain of Brat, a conserved structural motif present in a large family of growth regulators. In this report, we describe the crystal structure of the Brat NHL domain and present a model of the Pumilio-Brat complex derived from in silico docking experiments and supported by mutational analysis of the protein-protein interface.

RNA recognition via the SAM domain of Smaug.

The Nanos protein gradient in Drosophila, required for proper abdominal segmentation, is generated in part via translational repression of its mRNA by Smaug. We report here the crystal structure of the Smaug RNA binding domain, which shows no sequence homology to any previously characterized RNA binding motif. The structure reveals an unusual makeup in which a SAM domain, a common protein-protein interaction module, is affixed to a pseudo-HEAT repeat analogous topology (PHAT) domain.

Translational repressors in Drosophila.

Translational regulation is an important aspect of gene regulation, particularly during early development of the fruit fly embryo when transcriptional mechanisms are untenable. Study of pattern formation and dosage compensation has identified several repressors that bind discrete sites in the untranslated portions of target mRNAs. These repressors do not work in isolation - each binds multiple sites in the appropriate mRNA, and the resulting RNA-protein complexes appear to recruit co-repressors by a variety of mechanisms.

Crystallization and characterization of Smaug: a novel RNA-binding motif.

During Drosophila embryogenesis, Smaug protein represses translation of Nanos through an interaction with a specific element in its 3(')UTR. The repression occurs in the bulk cytoplasm of the embryo; Nanos is, however, successfully translated in the specialized cytoplasm of the posterior pole. This generates a gradient of Nanos emanating from the posterior pole that is essential for organizing proper abdominal segmentation.