scholarly journals The FMRP–MOV10 complex: a translational regulatory switch modulated by G-Quadruplexes

2019 ◽  
Author(s):  
Phillip J Kenny ◽  
Miri Kim ◽  
Geena Skariah ◽  
Joshua Nielsen ◽  
Monica C Lannom ◽  
...  
Keyword(s):  
Translational Regulation ◽  
Rna Binding ◽  
Fragile X ◽  
Leukemia Virus ◽  
Over Expression ◽  
G Quadruplex ◽  
N Terminus ◽  
Moloney Leukemia Virus ◽  
Endogenous Proteins ◽  
Normal Cognition

Abstract The Fragile X Mental Retardation Protein (FMRP) is an RNA binding protein that regulates translation and is required for normal cognition. FMRP upregulates and downregulates the activity of microRNA (miRNA)-mediated silencing in the 3′ UTR of a subset of mRNAs through its interaction with RNA helicase Moloney leukemia virus 10 (MOV10). This bi-functional role is modulated through RNA secondary structures known as G-Quadruplexes. We elucidated the mechanism of FMRP’s role in suppressing Argonaute (AGO) family members’ association with mRNAs by mapping the interacting domains of FMRP, MOV10 and AGO and then showed that the RGG box of FMRP protects a subset of co-bound mRNAs from AGO association. The N-terminus of MOV10 is required for this protection: its over-expression leads to increased levels of the endogenous proteins encoded by this co-bound subset of mRNAs. The N-terminus of MOV10 also leads to increased RGG box-dependent binding to the SC1 RNA G-Quadruplex and is required for outgrowth of neurites. Lastly, we showed that FMRP has a global role in miRNA-mediated translational regulation by recruiting AGO2 to a large subset of RNAs in mouse brain.

scholarly journals Crystal structure reveals specific recognition of a G-quadruplex RNA by a β-turn in the RGG motif of FMRP

2015 ◽  
Vol 112 (39) ◽  
pp. E5391-E5400 ◽  
Author(s):  
Nikita Vasilyev ◽  
Anna Polonskaia ◽  
Jennifer C. Darnell ◽  
Robert B. Darnell ◽  
Dinshaw J. Patel ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Rna Binding ◽  
Specific Binding ◽  
Fragile X ◽  
Type I ◽  
Shape Complementarity ◽  
G Quadruplex ◽  
Rna Duplexes ◽  
Human Disorders

Fragile X Mental Retardation Protein (FMRP) is a regulatory RNA binding protein that plays a central role in the development of several human disorders including Fragile X Syndrome (FXS) and autism. FMRP uses an arginine-glycine-rich (RGG) motif for specific interactions with guanine (G)-quadruplexes, mRNA elements implicated in the disease-associated regulation of specific mRNAs. Here we report the 2.8-Å crystal structure of the complex between the human FMRP RGG peptide bound to the in vitro selected G-rich RNA. In this model system, the RNA adopts an intramolecular K+-stabilized G-quadruplex structure composed of three G-quartets and a mixed tetrad connected to an RNA duplex. The RGG peptide specifically binds to the duplex–quadruplex junction, the mixed tetrad, and the duplex region of the RNA through shape complementarity, cation–π interactions, and multiple hydrogen bonds. Many of these interactions critically depend on a type I β-turn, a secondary structure element whose formation was not previously recognized in the RGG motif of FMRP. RNA mutagenesis and footprinting experiments indicate that interactions of the peptide with the duplex–quadruplex junction and the duplex of RNA are equally important for affinity and specificity of the RGG–RNA complex formation. These results suggest that specific binding of cellular RNAs by FMRP may involve hydrogen bonding with RNA duplexes and that RNA duplex recognition can be a characteristic RNA binding feature for RGG motifs in other proteins.

scholarly journals FMRP promotes RNA localization to neuronal projections through interactions between its RGG domain and G-quadruplex RNA sequences

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Raeann Goering ◽  
Laura I Hudish ◽  
Bryan B Guzman ◽  
Nisha Raj ◽  
Gary J Bassell ◽  
...  
Keyword(s):  
Fragile X Syndrome ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Rna Localization ◽  
Null Mouse ◽  
Rna Sequences ◽  
Rna Molecules ◽  
G Quadruplex

The sorting of RNA molecules to subcellular locations facilitates the activity of spatially restricted processes. We have analyzed subcellular transcriptomes of FMRP-null mouse neuronal cells to identify transcripts that depend on FMRP for efficient transport to neurites. We found that these transcripts contain an enrichment of G-quadruplex sequences in their 3′ UTRs, suggesting that FMRP recognizes them to promote RNA localization. We observed similar results in neurons derived from Fragile X Syndrome patients. We identified the RGG domain of FMRP as important for binding G-quadruplexes and the transport of G-quadruplex-containing transcripts. Finally, we found that the translation and localization targets of FMRP were distinct and that an FMRP mutant that is unable to bind ribosomes still promoted localization of G-quadruplex-containing messages. This suggests that these two regulatory modes of FMRP may be functionally separated. These results provide a framework for the elucidation of similar mechanisms governed by other RNA-binding proteins.

The N-Terminus of the Fragile X Mental Retardation Protein Contains a Novel Domain Involved in Dimerization and RNA Binding†

Biochemistry ◽  
2003 ◽  
Vol 42 (35) ◽  
pp. 10437-10444 ◽  
Author(s):  
S. Adinolfi ◽  
A. Ramos ◽  
S. R. Martin ◽  
F. Dal Piaz ◽  
P. Pucci ◽  
...  
Keyword(s):  
Mental Retardation ◽  
Rna Binding ◽  
Fragile X ◽  
Fragile X Mental Retardation ◽  
Mental Retardation Protein ◽  
N Terminus

scholarly journals Effects of Moloney Leukemia Virus 10 Protein on Hepatitis B Virus Infection and Viral Replication

Viruses ◽  
2019 ◽  
Vol 11 (7) ◽  
pp. 651 ◽  
Author(s):  
Maritza Puray-Chavez ◽  
Mahmoud Farghali ◽  
Vincent Yapo ◽  
Andrew Huber ◽  
Dandan Liu ◽  
...  
Keyword(s):  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Cell Lines ◽  
Core Protein ◽  
Leukemia Virus ◽  
Hbv Dna ◽  
Genomic Rna ◽  
Over Expression ◽  
B Virus ◽  
Moloney Leukemia Virus

Moloney leukemia virus 10 (MOV10) is an RNA helicase that has been shown to affect the replication of several viruses. The effect of MOV10 on Hepatitis B virus (HBV) infection is not known and its role on the replication of this virus is poorly understood. We investigated the effect of MOV10 down-regulation and MOV10 over-expression on HBV in a variety of cell lines, as well as in an infection system using a replication competent virus. We report that MOV10 down-regulation, using siRNA, shRNA, and CRISPR/Cas9 gene editing technology, resulted in increased levels of HBV DNA, HBV pre-genomic RNA, and HBV core protein. In contrast, MOV10 over-expression reduced HBV DNA, HBV pre-genomic RNA, and HBV core protein. These effects were consistent in all tested cell lines, providing strong evidence for the involvement of MOV10 in the HBV life cycle. We demonstrated that MOV10 does not interact with HBV-core. However, MOV10 binds HBV pgRNA and this interaction does not affect HBV pgRNA decay rate. We conclude that the restriction of HBV by MOV10 is mediated through effects at the level of viral RNA.

scholarly journals Dynamic Association of the Fragile X Mental Retardation Protein as a Messenger Ribonucleoprotein between Microtubules and Polyribosomes

2008 ◽  
Vol 19 (1) ◽  
pp. 105-114 ◽  
Author(s):  
Houping Wang ◽  
Jason B. Dictenberg ◽  
Li Ku ◽  
Wen Li ◽  
Gary J. Bassell ◽  
...  
Keyword(s):  
Mental Retardation ◽  
Translational Regulation ◽  
Rna Binding ◽  
Fragile X ◽  
Synaptic Function ◽  
Dependent Manner ◽  
Fragile X Mental Retardation ◽  
Messenger Ribonucleoprotein ◽  
Mental Retardation Protein ◽  
Neuronal Processes

The fragile X mental retardation protein (FMRP) is a selective RNA-binding protein that regulates translation and plays essential roles in synaptic function. FMRP is bound to specific mRNA ligands, actively transported into neuronal processes in a microtubule-dependent manner, and associated with polyribosomes engaged in translation elongation. However, the biochemical relationship between FMRP–microtubule association and FMRP–polyribosome association remains elusive. Here, we report that although the majority of FMRP is incorporated into elongating polyribosomes in the soluble cytoplasm, microtubule-associated FMRP is predominantly retained in translationally dormant, polyribosome-free messenger ribonucleoprotein (mRNP) complexes. Interestingly, FMRP–microtubule association is increased when mRNPs are dynamically released from polyribosomes as a result of inhibiting translation initiation. Furthermore, the I304N mutant FMRP that fails to be incorporated into polyribosomes is associated with microtubules in mRNP particles and transported into neuronal dendrites in a microtubule-dependent, 3,5-dihydroxyphenylglycine-stimulated manner with similar kinetics to that of wild-type FMRP. Hence, polyribosome-free FMRP–mRNP complexes travel on microtubules and wait for activity-dependent translational derepression at the site of function. The dual participation of FMRP in dormant mRNPs and polyribosomes suggests distinct roles of FMRP in dendritic transport and translational regulation, two distinct phases that control local protein production to accommodate synaptic plasticity.

scholarly journals FMRP and MOV10 regulate Dicer1 expression and dendrite development

PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0260005
Author(s):  
Monica C. Lannom ◽  
Joshua Nielsen ◽  
Aatiqa Nawaz ◽  
Temirlan Shilikbay ◽  
Stephanie Ceman
Keyword(s):  
Fragile X ◽  
Leukemia Virus ◽  
Dendritic Morphology ◽  
Neuronal Morphology ◽  
Translation Regulation ◽  
Neuro2a Cell ◽  
Dendrite Development ◽  
Moloney Leukemia Virus ◽  
Dicer Expression ◽  
Dicer1 Expression

Fragile X syndrome results from the loss of expression of the Fragile X Mental Retardation Protein (FMRP). FMRP and RNA helicase Moloney Leukemia virus 10 (MOV10) are important Argonaute (AGO) cofactors for miRNA-mediated translation regulation. We previously showed that MOV10 functionally associates with FMRP. Here we quantify the effect of reduced MOV10 and FMRP expression on dendritic morphology. Murine neurons with reduced MOV10 and FMRP phenocopied Dicer1 KO neurons which exhibit impaired dendritic maturation Hong J (2013), leading us to hypothesize that MOV10 and FMRP regulate DICER expression. In cells and tissues expressing reduced MOV10 or no FMRP, DICER expression was significantly reduced. Moreover, the Dicer1 mRNA is a Cross-Linking Immunoprecipitation (CLIP) target of FMRP Darnell JC (2011), MOV10 Skariah G (2017) and AGO2 Kenny PJ (2020). MOV10 and FMRP modulate expression of DICER1 mRNA through its 3’untranslated region (UTR) and introduction of a DICER1 transgene restores normal neurite outgrowth in the Mov10 KO neuroblastoma Neuro2A cell line and branching in MOV10 heterozygote neurons. Moreover, we observe a global reduction in AGO2-associated microRNAs isolated from Fmr1 KO brain. We conclude that the MOV10-FMRP-AGO2 complex regulates DICER expression, revealing a novel mechanism for regulation of miRNA production required for normal neuronal morphology.

scholarly journals FMRP promotes RNA localization to neuronal projections through interactions between its RGG domain and G-quadruplex RNA sequences

2019 ◽  
Author(s):  
Raeann Goering ◽  
Laura I. Hudish ◽  
Bryan B. Guzman ◽  
Nisha Raj ◽  
Gary J. Bassell ◽  
...  
Keyword(s):  
Single Molecule ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Fragile X ◽  
Rna Localization ◽  
Null Mouse ◽  
Rna Sequences ◽  
Local Protein Synthesis ◽  
Rna Molecules ◽  
G Quadruplex

ABSTRACTThe sorting of RNA molecules to distinct subcellular locations facilitates the activity of spatially restricted processes through local protein synthesis. This process affects thousands of transcripts yet precisely how these RNAs are trafficked to their destinations remains generally unclear. Here we have analyzed subcellular transcriptomes of FMRP-null mouse neuronal cells to identify transcripts that depend on FMRP for efficient transport to neurites. We found that these FMRP RNA localization targets contain a large enrichment of G-quadruplex sequences, particularly in their 3′ UTRs, suggesting that FMRP recognizes these sequences to promote the localization of transcripts that contain them. Fractionation of neurons derived from human Fragile X Syndrome patients revealed a high degree of conservation in the identity of FMRP localization targets between human and mouse as well as an enrichment of G-quadruplex sequences in human FMRP RNA localization targets. Using high-throughput RNA/protein interaction assays and single-molecule RNA FISH, we identified the RGG domain of FMRP as important for both interaction with G-quadruplex RNA sequences and the neuronal transport of G-quadruplex-containing transcripts. Finally, we used ribosome footprinting to identify translational regulatory targets of FMRP. The translational regulatory targets were not enriched for G-quadruplex sequences and were largely distinct from the RNA localization targets of FMRP, indicating that the two functions can be biochemically separated and are mediated through different target recognition mechanisms. These results establish a molecular mechanism underlying FMRP-mediated neuronal RNA localization and provide a framework for the elucidation of similar mechanisms governed by other RNA-binding proteins.

scholarly journals Heterogeneous Nuclear Ribonucleoproteins: Implications in Neurological Diseases

2020 ◽  
Author(s):  
Yi-Hua Low ◽  
Yasmine Asi ◽  
Sandrine C. Foti ◽  
Tammaryn Lashley
Keyword(s):  
Translational Regulation ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Muscular Atrophy ◽  
Fragile X ◽  
Neurological Diseases ◽  
Congenital Myasthenic Syndrome ◽  
Granule Formation ◽  
Myasthenic Syndrome ◽  
Ataxia Syndrome

Abstract Heterogenous nuclear ribonucleoproteins (hnRNPs) are a complex and functionally diverse family of RNA binding proteins with multifarious roles. They are involved, directly or indirectly, in alternative splicing, transcriptional and translational regulation, stress granule formation, cell cycle regulation, and axonal transport. It is unsurprising, given their heavy involvement in maintaining functional integrity of the cell, that their dysfunction has neurological implications. However, compared to their more established roles in cancer, the evidence of hnRNP implication in neurological diseases is still in its infancy. This review aims to consolidate the evidences for hnRNP involvement in neurological diseases, with a focus on spinal muscular atrophy (SMA), Alzheimer’s disease (AD), amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), multiple sclerosis (MS), congenital myasthenic syndrome (CMS), and fragile X-associated tremor/ataxia syndrome (FXTAS). Understanding more about hnRNP involvement in neurological diseases can further elucidate the pathomechanisms involved in these diseases and perhaps guide future therapeutic advances.

scholarly journals Independent role for presynaptic FMRP revealed by an FMR1 missense mutation associated with intellectual disability and seizures

2015 ◽  
Vol 112 (4) ◽  
pp. 949-956 ◽  
Author(s):  
Leila K. Myrick ◽  
Pan-Yue Deng ◽  
Hideharu Hashimoto ◽  
Young Mi Oh ◽  
Yongcheol Cho ◽  
...  
Keyword(s):  
Mental Retardation ◽  
Intellectual Disability ◽  
Missense Mutation ◽  
Translational Regulation ◽  
Rna Binding ◽  
Fragile X ◽  
Bk Channels ◽  
Structural Defects ◽  
Fragile X Mental Retardation ◽  
Specific Subset

Fragile X syndrome (FXS) results in intellectual disability (ID) most often caused by silencing of the fragile X mental retardation 1 (FMR1) gene. The resulting absence of fragile X mental retardation protein 1 (FMRP) leads to both pre- and postsynaptic defects, yet whether the pre- and postsynaptic functions of FMRP are independent and have distinct roles in FXS neuropathology remain poorly understood. Here, we demonstrate an independent presynaptic function for FMRP through the study of an ID patient with an FMR1 missense mutation. This mutation, c.413G > A (R138Q), preserves FMRP’s canonical functions in RNA binding and translational regulation, which are traditionally associated with postsynaptic compartments. However, neuronally driven expression of the mutant FMRP is unable to rescue structural defects at the neuromuscular junction in fragile x mental retardation 1 (dfmr1)-deficient Drosophila, suggesting a presynaptic-specific impairment. Furthermore, mutant FMRP loses the ability to rescue presynaptic action potential (AP) broadening in Fmr1 KO mice. The R138Q mutation also disrupts FMRP’s interaction with the large-conductance calcium-activated potassium (BK) channels that modulate AP width. These results reveal a presynaptic- and translation-independent function of FMRP that is linked to a specific subset of FXS phenotypes.

scholarly journals NF90 Interacts with Components of RISC and Modulates Association of Ago2 with mRNA

2021 ◽  
Author(s):  
Giuseppa Grasso ◽  
Celine Franckhauser ◽  
Rima Nait-Saidi ◽  
Maxime Bello ◽  
Jerome Barbier ◽  
...  
Keyword(s):  
Rna Binding ◽  
Leukemia Virus ◽  
Vascular Endothelial ◽  
Cellular Mechanisms ◽  
Vegf Mrna ◽  
Argonaute 2 ◽  
Target Mrnas ◽  
Moloney Leukemia Virus ◽  
Endothelial Growth Factor ◽  
Common Target

Nuclear Factor 90 (NF90) is a double-stranded RNA-binding protein involved in a multitude of different cellular mechanisms such as transcription, translation, viral infection and mRNA stability. Recent data suggest that NF90 might influence the abundance of target mRNAs in the cytoplasm through miRNA- and Argonaute 2 (Ago2)-dependent activity. Here, we identified the interactome of NF90 in the cytoplasm, which revealed several components of the RNA-induced silencing complex (RISC) and associated factors. Co-immunoprecipitation analysis confirmed the interaction of NF90 with the RISC-associated RNA helicase, Moloney leukemia virus 10 (MOV10), and other proteins involved in RISC-mediated silencing, including Ago2. Furthermore, NF90 association with MOV10 and Ago2 was found to be RNA-dependent. Glycerol gradient sedimentation of NF90 immune complexes indicates that these proteins occur in the same protein complex. At target RNAs predicted to bind both NF90 and MOV10 in their 3' UTRs, NF90 association was increased upon loss of MOV10 and vice versa, suggesting that the two proteins may compete for the binding of common target mRNAs. Interestingly, loss of NF90 led to an increase in association of Ago2 as well as a decrease in the abundance of the target mRNA. Similarly, during hypoxia, the binding of Ago2 to vascular endothelial growth factor (VEGF) mRNA increased after loss of NF90, while the level of VEGF mRNA decreased. These findings suggest a role for NF90 in the regulation of RISC-mediated silencing which stabilizes target mRNAs, such as VEGF, during cancer-induced hypoxia.

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