Fragile X syndrome is a leading heritable cause of mental retardation that results from the loss of FMR1 gene function. A Drosophila model for Fragile X syndrome, based on the loss of dfmr1 activity, exhibits phenotypes that bear similarity to Fragile X-related symptoms. Herein, we demonstrate that treatment with metabotropic glutamate receptor (mGluR) antagonists or lithium can rescue courtship and mushroom body defects observed in these flies. Furthermore, we demonstrate that dfmr1 mutants display cognitive deficits in experience-dependent modification of courtship behavior, and treatment with mGluR antagonists or lithium restores these memory defects. These findings implicate enhanced mGluR signaling as the underlying cause of the cognitive, as well as some of the behavioral and neuronal, phenotypes observed in the Drosophila Fragile X model. They also raise the possibility that compounds having similar effects on metabotropic glutamate receptors may ameliorate cognitive and behavioral defects observed in Fragile X patients.

Fragile X syndrome is caused by a loss of expression of the fragile X mental retardation protein (FMRP). FMRP is a selective RNA-binding protein which forms a messenger ribonucleoprotein (mRNP) complex that associates with polyribosomes. Recently, mRNA ligands associated with FMRP have been identified. However, the mechanism by which FMRP regulates the translation of its mRNA ligands remains unclear. MicroRNAs are small noncoding RNAs involved in translational control. Here we show that in vivo mammalian FMRP interacts with microRNAs and the components of the microRNA pathways including Dicer and the mammalian ortholog of Argonaute 1 (AGO1). Using two different Drosophila melanogaster models, we show that AGO1 is critical for FMRP function in neural development and synaptogenesis. Our results suggest that FMRP may regulate neuronal translation via microRNAs and links microRNAs with human disease.

Fragile X mental retardation is a prominent genetic disorder caused by the lack of the FMR1 gene product, a known RNA binding protein. Specific physiologic pathways regulated by FMR1 function have yet to be identified. Adult dfmr1 (also called dfxr) mutant flies display arrhythmic circadian activity and have erratic patterns of locomotor activity, whereas overexpression of dFMR1 leads to a lengthened period. dfmr1 mutant males also display reduced courtship activity which appears to result from their inability to maintain courtship interest. Molecular analysis fails to reveal any defects in the expression of clock components; however, the CREB output is affected. Morphological analysis of neurons required for normal circadian behavior reveals subtle abnormalities, suggesting that defects in axonal pathfinding or synapse formation may cause the observed behavioral defects.

Fragile X syndrome is the most common inherited form of mental retardation. It is caused by loss of FMR1 gene activity due to either lack of expression or expression of a mutant form of the protein. In mammals, FMR1 is a member of a small protein family that consists of FMR1, FXR1, and FXR2. All three members bind RNA and contain sequence motifs that are commonly found in RNA-binding proteins, including two KH domains and an RGG box. The FMR1/FXR proteins also contain a 60S ribosomal subunit interaction domain and a protein-protein interaction domain which mediates homomer and heteromer formation with each family member. Nevertheless, the specific molecular functions of FMR1/FXR proteins are unknown. Here we report the cloning and characterization of a Drosophila melanogaster homolog of the mammalian FMR1/FXR gene family. This first invertebrate homolog, termed dfmr1, has a high degree of amino acid sequence identity/ similarity with the defined functional domains of the FMR1/FXR proteins. The dfmr1 product binds RNA and is similar in subcellular localization and embryonic expression pattern to the mammalian FMR1/FXR proteins. Overexpression of dfmr1 driven by the UAS-GAL4 system leads to apoptotic cell loss in all adult Drosophila tissues examined. This phenotype is dependent on the activity of the KH domains. The ability to induce a dominant phenotype by overexpressing dfmr1 opens the possibility of using genetic approaches in Drosophila to identify the pathways in which the FMR1/FXR proteins function.Fragile X syndrome is the most common form of hereditary mental retardation whose effects are traced to the loss of function of a single gene, named FMR1. This syndrome affects approximately 1 in 5,000 male births and is globally distributed throughout the human population (19,42,61). In most cases, the disease results from the repression of FMR1 gene expression that is due to an expansion of a CGG trinucleotide repeat in the 5Ј untranslated region of the gene (25,28,36,45,65,66,73). Subsequent methylation of this expanded repeat results in transcriptional silencing of the FMR1 gene (7, 43). A few fragile X patients with partial or complete deletions of the FMR1 gene have been identified, and these patients have phenotypes similar to those affected by the trinucleotide repeat expansion (26, 68). One patient who has a single point mutation in the FMR1 gene that replaces an isoleucine residue at amino acid 304 with asparagine (I304N) exhibits a particularly severe fragile X phenotype (17). The severity of the phenotype observed in this patient has prompted the suggestion that the I304N substitution results in a dominant-negative form of FMR1 protein (22, 53).The FMR1 protein binds RNA in vitro and contains two types of RNA-binding motifs, KH domains and an RGG box (10, 35, 55). The RNA-binding activity of FMR1 appears to be selective. It has been estimated that FMR1 interacts with about 4% of human fetal brain mRNAs, including its own mRNA (4). The importance of the RNA-binding activity to the function of the FMR1 ...

Piwi family proteins are essential for germline development and bind piwi-interacting RNAs (piRNAs 1 2 3). The grandchildless gene aub of Drosophila melanogaster encodes the piRNA-binding protein Aub that is essential for formation of primordial germ cells (PGCs) 4. Here we report that mouse, Xenopus laevis and Drosophila melanogaster Piwi family proteins contain symmetrical dimethylarginines (sDMAs). We find that Piwi proteins are expressed in X. laevis oocytes and we identify numerous X. laevis piRNAs. We report that the Drosophila homolog of protein methyltransferase 5 (dPRMT5, csul/dart5), which is also the product of a grandchildless gene 5, 6, is required for arginine methylation of Drosophila Piwi, Ago3 and Aub proteins, in vivo. Loss of dPRMT5 activity leads to reduction of piRNAs and in particular of Ago3 and Aub protein levels and accumulation of retrotransposons in the Drosophila ovary. Our studies explain the relationship between aub and dPRMT5 (csul/dart5) genes by demonstrating that dPRMT5 is the enzyme that methylates Aub. Our findings underscore the significance of sDMA modification of Piwi proteins in the germline and suggest an interacting pathway of genes that are required for piRNA function and PGC specification.