A 3' untranslated region variant in FMR1 eliminates neuronal activity-dependent translation of FMRP by disrupting binding of the RNA-binding protein HuR (original) (raw)
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Biology of the fragile X mental retardation protein, an RNA-binding protein
Biochemistry and Cell Biology, 1999
The fragile X syndrome, an X-linked disease, is the most frequent cause of inherited mental retardation. The syndrome results from the absence of expression of the FMR1 gene (fragile mental retardation 1) owing to the expansion of a CGG trinucleotide repeat located in the 5′ untranslated region of the gene and the subsequent methylation of its CpG island. The FMR1 gene product (FMRP) is a cytoplasmic protein that contains two KH domains and one RGG box, characteristics of RNA-binding proteins. FMRP is associated with mRNP complexes containing poly(A)+mRNA within actively translating polyribosomes and contains nuclear localization and export signals making it a putative transporter (chaperone) of mRNA from the nucleus to the cytoplasm. FMRP is the archetype of a novel family of cytoplasmic RNA-binding proteins that includes FXR1P and FXR2P. Both of these proteins are very similar in overall structure to FMRP and are also associated with cytoplasmic mRNPs. Members of the FMR family are widely expressed in mouse and human tissues, albeit at various levels, and seem to play a subtle choreography of expression. FMRP is most abundant in neurons and is absent in muscle. FXR1P is strongly expressed in muscle and low levels are detected in neurons. The complex expression patterns of the FMR1 gene family in different cells and tissues suggest that independent, however similar, functions for each of the three FMR-related proteins might be expected in the selection and metabolism of tissue-specific classes of mRNA. The molecular mechanisms altered in cells lacking FMRP still remain to be elucidated as well as the putative role(s) of FXR1P and FXR2P as compensatory molecules.
Fragile X Mental Retardation Protein Targets G Quartet mRNAs Important for Neuronal Function
Cell, 2001
RNA secondary structure, thereby allowing access to specific sequences by other RNA binding domains (Ghi-and Pediatrics Emory University School of Medicine solfi et al., 1992a, 1992b; Kiledjian and Dreyfuss, 1992). Although sequence-specific RNA-protein interactions Atlanta, Georgia 30322 have been defined for several KH-type RNA binding proteins including the neuronal paraneoplastic disease antigen Nova (Buckanovich and Darnell, 1997; Jensen Summary et al., 2000b), the splicing factor SF1/BBP (Berglund et al., 1998), and hnRNPs K and E1/E2 (Ostareck et al., Loss of fragile X mental retardation protein (FMRP) function causes the fragile X mental retardation syn-1997; Thisted et al., 2001), the RNA specificity with which FMRP binds RNA has remained unexplored. This ques-drome. FMRP harbors three RNA binding domains, associates with polysomes, and is thought to regulate tion is of particular interest to the fragile X syndrome, since one severely affected individual harbors a single mRNA translation and/or localization, but the RNAs to which it binds are unknown. We have used RNA amino acid change (I304N) within the second KH domain of FMRP (DeBoulle et al., 1993). Some of the conse-selection to demonstrate that the FMRP RGG box binds intramolecular G quartets. This data allowed us quences of this mutation on FMRP, such as its inability to form a homodimer (Absher et al., 2001; Laggerbauer to identify mRNAs encoding proteins involved in synaptic or developmental neurobiology that harbor et al., 2001), failure of the mutant protein to interact with polyribosomes (Absher et al., 2001; Feng et al., 1997a), FMRP binding elements. The majority of these mRNAs have an altered polysome association in fragile X pa-and more rapid protein shuttling between the nucleocytoplasmic space (Tamanini et al., 1999), have been tient cells. These data demonstrate that G quartets serve as physiologically relevant targets for FMRP and noted. Cocrystallization of the Nova KH3 domain with its RNA target predicts that the mutated isoleucine in identify mRNAs whose dysregulation may underlie human mental retardation. FMRP (I304N) lies at the core of the hydrophobic RNA binding pocket of the KH domain (Lewis et al . These observations suggest that identification of the Introduction RNA targets to which FMRP binds will yield biologically important insight into its function. The fragile X syndrome is the most common form of inherited mental retardation, manifested by mild to mod-The role of FMRP as an RNA binding protein has been most clearly established by the demonstration that erate cognitive and behavioral abnormalities accompanied by macroorchidism and subtle craniofacial dysmor-%58ف of cellular FMRP is present on actively translating polyribosomes (Feng et al., 1997a). Interestingly, the phia. The syndrome usually results from the expansion and hypermethylation of a CGG repeat in the 5Ј UTR of I304N mutant FMRP is not found in the polysome fractions (Feng et al., 1997a), consistent with an important the FMR1 gene that results in transcriptional silencing (reviewed by Jin and Warren, 2000), indicating that the role for the protein in regulating mRNA translation. Within the brain, FMRP is highly concentrated in neurons presence of FMRP is essential for higher cognitive function. with little or no expression in glia (Abitbol et al., 1993; Cloning the FMR1 gene led to the finding that FMRP Devys et al., 1993; Hinds et al., 1993). At a subcellular is an RNA binding protein (Ashley et al., 1993; Siomi et level, FMRP localizes to the perikaryon and dendrites al., 1993b). FMRP contains two tandem RNA binding in a pattern consistent with the location of neuronal KH domains and a C-terminal RGG box. KH domains polysomes (Feng et al., 1997b). It has both a nuclear were first described in hnRNP K (Siomi et al., 1993a) localization signal and a Rev-like export signal and is and subsequently were found to be present in a number believed to shuttle in and out of the nucleus (Eberhart of RNA binding proteins of diverse function, including et al., 1996; Feng et al., 1997b; Fridell et al., 1996; Sittler RNA localization (zip code binding protein), translational et al., 1996). The most profound finding on autopsy in control (hnRNP K and hnRNPE1/E2), pre-mRNA splicing FMRP patients is the presence of very long and thin (Nova-1, MER-1, SF1, KSRP, and PSI), shuttling trandendritic spines in the neocortex (Hinton et al., 1991; scripts to the cytoplasm (hnRNP K), and regulation of Rudelli et al., 1985), and similar dendritic spine abnor-mRNA stability (hnRNP E1/E2) (reviewed by Burd and malities are present in Fmr1 null mice (Comery et al., Dreyfuss, 1994; Lewis et al., 2000). RGG boxes are found 1997; Nimchinsky et al., 2001). One unifying model for FMRP function is that it shuttles mRNAs from the nucleus to postsynaptic sites where mRNAs are held in a
Nucleic Acids Research, 2006
Fragile X syndrome, the most frequent form of inherited mental retardation, is due to the absence of expression of the Fragile X Mental Retardation Protein (FMRP), an RNA binding protein with high specificity for G-quartet RNA structure. FMRP is involved in several steps of mRNA metabolism: nucleocytoplasmic trafficking, translational control and transport along dendrites in neurons. Fragile X Related Protein 1 (FXR1P), a homologue and interactor of FMRP, has been postulated to have a function similar to FMRP, leading to the hypothesis that it can compensate for the absence of FMRP in Fragile X patients. Here we analyze the ability of three isoforms of FXR1P, expressed in different tissues, to bind G-quartet RNA structure specifically. Only the longest FXR1P isoform was found to be able to bind specifically the G-quartet RNA, albeit with a lower affinity as compared to FMRP, whereas the other two isoforms negatively regulate the affinity of FMRP for G-quartet RNA. This result is important to decipher the molecular basis of fragile X syndrome, through the understanding of FMRP action in the context of its multimolecular complex in different tissues. In addition, we show that the action of FXR1P is synergistic rather than compensatory for FMRP function.
Cell, 2001
genes FXR1 and FXR2, encode a small family of RNA binding proteins that share over 60% amino acid identity (Ashley et al., 1993;. As an RNA binding protein, FMRP has been shown to bind to RNA homopolymers as well as a subset of brain transcripts in vitro (Ashley et al., 1993;). complex is composed of several proteins including other RNA binding proteins such as FXR1P and FXR2P (Bar-Atlanta, Georgia 30322 4 Laboratory of Molecular Neuro-Oncology doni et al., 1999; Ceman et al., 1999, 2000; Schenck et al., 2001). Purified FMRP displays an intrinsic RNA The Rockefeller University New York, New York 10021 binding capacity (Brown et al., 1998), and in the cytoplasm, FMRP-mRNP is associated with translating polymutation in the second KH domain of FMRP (I304N), which results in a severe fragile X phenotype, prevents this polyribosome association, suggesting that the Summary FMRP association with polyribosomes is functionally important (De Boulle et al., 1993; Feng et al., 1997a). Fragile X syndrome results from the absence of the RNA binding FMR protein. Here, mRNA was coimmu-
RNA-Binding Specificity of the Human Fragile X Mental Retardation Protein
Journal of Molecular Biology, 2020
Fragile X syndrome (FXS) is the most common form of inherited intellectual disability and is caused by a deficiency of the fragile X mental retardation protein (FMRP) in neurons. FMRP regulates the translation of numerous mRNAs within dendritic synapses, but how FMRP recognizes these target mRNAs remains unknown. FMRP has KH0, KH1, KH2, and RGG domains, which are thought to bind to specific RNA recognition elements (RREs). Several studies used high-throughput methods to identify various RREs in mRNAs that FMRP may bind to in vivo. However, there is little overlap in the mRNA targets identified by each study, suggesting that the RNA-binding specificity of FMRP is still unknown. To determine the specificity of FMRP for the RREs, we performed quantitative in vitro RNA binding studies with various constructs of human FMRP. Unexpectedly, our studies show that the KH domains do not bind to the previously identified RREs. To further investigate the RNA-binding specificity of FMRP, we developed a new method called Motif Identification by Analysis of Simple sequences (MIDAS) to identify singlestranded RNA (ssRNA) sequences bound by KH domains. We find that the FMRP KH0, KH1, and KH2 domains bind weakly to the ssRNA sequences suggesting that they may have evolved to bind more complex RNA structures. Additionally, we find that the RGG motif of human FMRP binds with a high affinity to an RNA G-quadruplex (GQ) structure that lacks single-stranded loops, double-stranded stems, or junctions.
The fragile X mental retardation protein binds specifically to its mRNA via a purine quartet motif
The EMBO Journal, 2001
Fragile X syndrome is caused by the absence of protein FMRP, the function of which is still poorly understood. Previous studies have suggested that FMRP may be involved in various aspects of mRNA metabolism, including transport, stability and/or translatability. FMRP was shown to interact with a subset of brain mRNAs as well as with its own mRNA; however, no speci®c RNA-binding site could be identi-®ed precisely. Here, we report the identi®cation and characterization of a speci®c and high af®nity binding site for FMRP in the RGG-coding region of its own mRNA. This site contains a purine quartet motif that is essential for FMRP binding and can be substituted by a heterologous quartet-forming motif. The speci®c binding of FMRP to its target site was con®rmed further in a reticulocyte lysate through its ability to repress translation of a reporter gene harboring the RNA target site in the 5¢-untranslated region. Our data address interesting questions concerning the role of FMRP in the post-transcriptional control of its own gene and possibly other target genes.
Journal of Neuroscience, 2008
Fragile X syndrome is the most common form of heritable mental retardation caused by the loss of function of the fragile X mental retardation protein FMRP. FMRP is a multidomain, RNA-binding protein involved in RNA transport and/or translational regulation. However, the binding specificity between FMRP and its various partners including interacting proteins and mRNA targets is essentially unknown. Previous work demonstrated that dFMRP, the Drosophila homolog of human FMRP, is structurally and functionally conserved with its mammalian counterparts. Here, we perform a forward genetic screen and isolate 26 missense mutations at 13 amino acid residues in the dFMRP coding dfmr1. Interestingly, all missense mutations identified affect highly conserved residues in the N terminal of dFMRP.
Nucleic acids research, 2018
Fragile X syndrome (FXS), the most common form of inherited intellectual disability, is due to the functional deficiency of the fragile X mental retardation protein (FMRP), an RNA-binding protein involved in translational regulation of many messenger RNAs, playing key roles in synaptic morphology and plasticity. To date, no effective treatment for FXS is available. We searched for FMRP targets by HITS-CLIP during early development of multiple mouse brain regions (hippocampus, cortex and cerebellum) at a time of brain development when FMRP is most highly expressed and synaptogenesis reaches a peak. We identified the largest dataset of mRNA targets of FMRP available in brain and we defined their cellular origin. We confirmed the G-quadruplex containing structure as an enriched motif in FMRP RNA targets. In addition to four less represented motifs, our study points out that, in the brain, CTGKA is the prominent motif bound by FMRP, which recognizes it when not engaged in Watson-Crick p...
From FMRP Function to Potential Therapies for Fragile X Syndrome
Neurochemical Research, 2013
Fragile X syndrome (FXS) is caused by mutations in the fragile X mental retardation 1 (FMR1) gene. Most FXS cases occur due to the expansion of the CGG trinucleotide repeats in the 5′ untranslated region (UTR) of FMR1, which leads to hypermethylation and in turn silences the expression of FMRP (fragile X mental retardation protein). Numerous studies have demonstrated that FMRP interacts with both coding and non-coding RNAs and represses protein synthesis at dendritic and synaptic locations. In the absence of FMRP, the basal protein translation is enhanced and not responsive to neuronal stimulation. The altered protein translation may contribute to functional abnormalities in certain aspects of synaptic plasticity and intracellular signaling triggered by Gq-coupled receptors. This review focuses on the current understanding of FMRP function and potential therapeutic strategies that are mainly based on the manipulation of FMRP targets and knowledge gained from FXS pathophysiology.