scholarly journals Identification of FMRP target mRNAs in the developmental brain: FMRP might coordinate Ras/MAPK, Wnt/β-catenin, and mTOR signaling during corticogenesis

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Cristine R. Casingal ◽  
Takako Kikkawa ◽  
Hitoshi Inada ◽  
Yukio Sasaki ◽  
Noriko Osumi
Keyword(s):  
Brain Development ◽  
Rna Binding ◽  
Fragile X ◽  
Autism Spectrum ◽  
Mtor Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Sequencing Analysis ◽  
Embryonic Brain ◽  
Target Mrnas ◽  
Mrna Targets

AbstractCorticogenesis is one of the most critical and complicated processes during embryonic brain development. Any slight impairment in corticogenesis could cause neurodevelopmental disorders such as Fragile X syndrome (FXS), of which symptoms contain intellectual disability (ID) and autism spectrum disorder (ASD). Fragile X mental retardation protein (FMRP), an RNA-binding protein responsible for FXS, shows strong expression in neural stem/precursor cells (NPCs) during corticogenesis, although its function during brain development remains largely unknown. In this study, we attempted to identify the FMRP target mRNAs in the cortical primordium using RNA immunoprecipitation sequencing analysis in the mouse embryonic brain. We identified 865 candidate genes as targets of FMRP involving 126 and 118 genes overlapped with ID and ASD-associated genes, respectively. These overlapped genes were enriched with those related to chromatin/chromosome organization and histone modifications, suggesting the involvement of FMRP in epigenetic regulation. We further identified a common set of 17 FMRP “core” target genes involved in neurogenesis/FXS/ID/ASD, containing factors associated with Ras/mitogen-activated protein kinase, Wnt/β-catenin, and mammalian target of rapamycin (mTOR) pathways. We indeed showed overactivation of mTOR signaling via an increase in mTOR phosphorylation in the Fmr1 knockout (Fmr1 KO) neocortex. Our results provide further insight into the critical roles of FMRP in the developing brain, where dysfunction of FMRP may influence the regulation of its mRNA targets affecting signaling pathways and epigenetic modifications.

scholarly journals Identification of FMRP target mRNAs in the developmental brain: FMRP might coordinate Ras/MAPK, Wnt/ß-catenin, and mTOR signaling during corticogenesis

2020 ◽  
Author(s):  
Cristine R. Casingal ◽  
Takako Kikkawa ◽  
Hitoshi Inada ◽  
Yukio Sasaki ◽  
Noriko Osumi
Keyword(s):  
Brain Development ◽  
Rna Binding ◽  
Fragile X ◽  
Autism Spectrum ◽  
Mtor Signaling ◽  
Mitogen Activated Protein Kinase ◽  
Sequencing Analysis ◽  
Embryonic Brain ◽  
Target Mrnas ◽  
Mrna Targets

Abstract Corticogenesis is one of the most critical and complicated processes during embryonic brain development. Any slight impairment in corticogenesis could cause neurodevelopmental disorders such as Fragile X syndrome (FXS), of which symptoms contain intellectual disability (ID) and autism spectrum disorder (ASD). Fragile X mental retardation protein (FMRP), an RNA-binding protein responsible for FXS, shows strong expression in neural stem/precursor cells (NPCs) during corticogenesis, although its function during brain development remains largely unknown. In this study, we attempted to identify the FMRP target mRNAs in the mouse cortical primordium using RNA immunoprecipitation sequencing analysis in the mouse embryonic brain. We identified 865 candidate genes as targets of FMRP involving 126 and 118 genes overlapped with ID and ASD-associated genes, respectively. These overlapped genes were enriched with those related to chromatin/chromosome organization and histone modifications, suggesting the involvement of FMRP in epigenetic regulation. We further identified a common set of 17 "core" genes involved in neurogenesis/FXS/ID/ASD, containing factors associated with Ras/mitogen-activated protein kinase, Wnt/ß-catenin, and mTOR pathways. We indeed showed overactivation of mTOR signaling via an increase in mTOR phosphorylation in the Fmr1 knockout (Fmr1 KO) neocortex. Our results provide further insight into the critical roles of FMRP in the developing brain, where dysfunction of FMRP may influence the regulation of its mRNA targets affecting signaling pathways and epigenetic modifications.

scholarly journals Identification of FMRP target genes expressed in corticogenesis: implication for common phenotypes among neurodevelopmental disorders

2019 ◽  
Author(s):  
Cristine R. Casingal ◽  
Takako Kikkawa ◽  
Hitoshi Inada ◽  
Noriko Osumi
Keyword(s):  
Transcriptional Control ◽  
Target Genes ◽  
Rna Binding ◽  
Fragile X ◽  
Autism Spectrum ◽  
Mrna Targets ◽  
Significant Difference ◽  
Rna Immunoprecipitation ◽  
The Brain

ABSTRACTFragile X mental retardation protein (FMRP) is encoded by FMR1 gene that is responsible for Fragile X Syndrome (FXS) showing intellectual disability and autism spectrum disorder. FMRP is an RNA binding protein highly expressed in the brain. Although several target genes for FMRP have been identified, limited studies have suggested the role of FMRP in corticogenesis. Here we performed RNA immunoprecipitation sequencing against the murine embryonic neocortex, and identified 124 genes as potential FMRP mRNA targets. We found 48 of these genes as overlapped with autism-related genes, which were categorized in four functional groups: “transcriptional regulation”, “regulation of actin cytoskeleton”, “ubiquitin-mediated proteolysis” and “calcium signaling pathway”. Four of these genes showed significant difference in expression in the cortical primordium of Fmr1-KO mice; Huwe1 and Kat6a increased, while Kmt2c and Apc decreased. Although the change in expression of these four genes was relatively small, these subtle changes due to dysregulated transcription could collectively contribute to impaired corticogenesis to cause phenotypes of FXS. Investigating the transcriptional control of FMRP on its mRNA targets may provide new insight to understand neurodevelopmental pathogenesis of FXS.

scholarly journals Star-PAP RNA Binding Landscape Reveals Novel Role of Star-PAP in mRNA Metabolism That Requires RBM10-RNA Association

2021 ◽  
Vol 22 (18) ◽  
pp. 9980
Author(s):  
Ganesh R. Koshre ◽  
Feba Shaji ◽  
Neeraja K. Mohanan ◽  
Nimmy Mohan ◽  
Jamshaid Ali ◽  
...  
Keyword(s):  
Rna Binding ◽  
Coding Region ◽  
High Association ◽  
Target Mrnas ◽  
Mrna Metabolism ◽  
Mrna Targets ◽  
Genome Wide ◽  
Global Profile ◽  
Specific Mrna

Star-PAP is a non-canonical poly(A) polymerase that selects mRNA targets for polyadenylation. Yet, genome-wide direct Star-PAP targets or the mechanism of specific mRNA recognition is still vague. Here, we employ HITS-CLIP to map the cellular Star-PAP binding landscape and the mechanism of global Star-PAP mRNA association. We show a transcriptome-wide association of Star-PAP that is diminished on Star-PAP depletion. Consistent with its role in the 3′-UTR processing, we observed a high association of Star-PAP at the 3′-UTR region. Strikingly, there is an enrichment of Star-PAP at the coding region exons (CDS) in 42% of target mRNAs. We demonstrate that Star-PAP binding de-stabilises these mRNAs indicating a new role of Star-PAP in mRNA metabolism. Comparison with earlier microarray data reveals that while UTR-associated transcripts are down-regulated, CDS-associated mRNAs are largely up-regulated on Star-PAP depletion. Strikingly, the knockdown of a Star-PAP coregulator RBM10 resulted in a global loss of Star-PAP association on target mRNAs. Consistently, RBM10 depletion compromises 3′-end processing of a set of Star-PAP target mRNAs, while regulating stability/turnover of a different set of mRNAs. Our results establish a global profile of Star-PAP mRNA association and a novel role of Star-PAP in the mRNA metabolism that requires RBM10-mRNA association in the cell.

scholarly journals Kinase pathway inhibition restores PSD95 induction in neurons lacking fragile X mental retardation protein

2019 ◽  
pp. 201812056
Author(s):  
Ying Yang ◽  
Yang Geng ◽  
Dongyun Jiang ◽  
Lin Ning ◽  
Hyung Joon Kim ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Mental Retardation ◽  
Rna Binding ◽  
Fragile X ◽  
Alternative Pathways ◽  
Fragile X Mental Retardation ◽  
Mrna Targets ◽  
Mental Retardation Protein ◽  
Pathway Inhibition

Fragile X syndrome (FXS) is the leading monogenic cause of autism and intellectual disability. FXS is caused by loss of expression of fragile X mental retardation protein (FMRP), an RNA-binding protein that regulates translation of numerous mRNA targets, some of which are present at synapses. While protein synthesis deficits have long been postulated as an etiology of FXS, how FMRP loss affects distributions of newly synthesized proteins is unknown. Here we investigated the role of FMRP in regulating expression of new copies of the synaptic protein PSD95 in an in vitro model of synaptic plasticity. We find that local BDNF application promotes persistent accumulation of new PSD95 at stimulated synapses and dendrites of cultured neurons, and that this accumulation is absent in FMRP-deficient mouse neurons. New PSD95 accumulation at sites of BDNF stimulation does not require known mechanisms regulating FMRP–mRNA interactions but instead requires the PI3K-mTORC1-S6K1 pathway. Surprisingly, in FMRP-deficient neurons, BDNF induction of new PSD95 accumulation can be restored by mTORC1-S6K1 blockade, suggesting that constitutively high mTORC1-S6K1 activity occludes PSD95 regulation by BDNF and that alternative pathways exist to mediate induction when mTORC1-S6K1 is inhibited. This study provides direct evidence for deficits in local protein synthesis and accumulation of newly synthesized protein in response to local stimulation in FXS, and supports mTORC1-S6K1 pathway inhibition as a potential therapeutic approach for FXS.

scholarly journals Disruption of the KH1 domain of Fmr1 leads to transcriptional alterations and attentional deficits in rats

2018 ◽  
Author(s):  
Carla E. M. Golden ◽  
Michael S. Breen ◽  
Lacin Koro ◽  
Sankalp Sonar ◽  
Kristi Niblo ◽  
...  
Keyword(s):  
Prefrontal Cortex ◽  
Medial Prefrontal Cortex ◽  
Rna Binding ◽  
Fragile X ◽  
Neurodevelopmental Disorder ◽  
Point Mutations ◽  
Autism Spectrum ◽  
Attention Deficits ◽  
Hub Genes ◽  
Transcriptional Profiles

AbstractFragile X Syndrome (FXS) is a neurodevelopmental disorder caused by mutations in the FMR1 gene. FXS is a leading monogenic cause of autism spectrum disorder (ASD) and inherited intellectual disability (ID). In most cases, the mutation is an expansion of a microsatellite (CGG triplet), which leads to suppressed expression of the fragile X mental retardation protein (FMRP), an RNA-binding protein involved in multiple aspects of mRNA metabolism. Interestingly, we found that the previously published Fmr1 knockout rat model of FXS expresses a transcript with an in-frame deletion of a K-homology (KH) domain, KH1. KH domains are RNA-binding domains of FMR1 and several of the few, known point mutations associated with FXS are found within them. We observed that this deletion leads to medial prefrontal cortex (mPFC)-dependent attention deficits, similar to those observed in FXS, and to alterations in transcriptional profiles within the mPFC, which mapped to two weighted gene coexpression network analysis modules. We demonstrated that these modules are conserved in human frontal cortex, are enriched for known FMRP targets and for genes involved in neuronal and synaptic processes, and that one is enriched for genes that are implicated in ASD, ID, and schizophrenia. Hub genes in these conserved modules represent potential targets for FXS. These findings provide support for a prefrontal deficit in FXS, indicate that attentional testing might be a reliable cross-species tool for investigating the pathophysiology of FXS and a potential readout for pharmacotherapy testing, and identify dysregulated gene expression modules in a relevant brain region.Significance StatementThe significance of the current study lies in two key domains. First, this study demonstrates that deletion of the Fmrp-KH1 domain is sufficient to cause major mPFC-dependent attention deficits in both males and females, like those observed in both individuals with FXS and in knockout mouse models for FXS. Second, the study shows that deletion of the KH1 domain leads to alterations in the transcriptional profiles within the medial prefrontal cortex (mPFC), which are of potential translational value for subjects with FXS. These findings indicate that attentional testing might be a reliable cross-species tool for investigating the pathophysiology of FXS and a potential readout for pharmacotherapy testing and also highlight hub genes for follow up.

scholarly journals Identification of TIA1 mRNA targets during human neuronal development

2021 ◽  
Author(s):  
Loryn P. Byres ◽  
Marat Mufteev ◽  
Kyoko E. Yuki ◽  
Wei Wei ◽  
Alina Piekna ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Neural Progenitor Cells ◽  
Neuronal Development ◽  
Rna Binding ◽  
Embryonic Stem ◽  
Autism Spectrum ◽  
Protein Levels ◽  
Mrna Targets ◽  
Autism Spectrum Disorder Risk ◽  
Rna Immunoprecipitation

Abstract Background Neuronal development is a tightly controlled process involving multi-layered regulatory mechanisms. While transcriptional pathways regulating neurodevelopment are well characterized, post-transcriptional programs are still poorly understood. TIA1 is an RNA-binding protein that can regulate splicing, stability, or translation of target mRNAs, and has been shown to play critical roles in stress response and neurodevelopment. However, the identity of mRNAs regulated by TIA1 during neurodevelopment under unstressed conditions is still unknown. Methods and Results To identify the mRNAs targeted by TIA1 during the first stages of human neurodevelopment, we performed RNA immunoprecipitation-sequencing (RIP-seq) on human embryonic stem cells (hESCs) and derived neural progenitor cells (NPCs), and cortical neurons under unstressed conditions. While there was no change in TIA1 protein levels, the number of TIA1 targeted mRNAs decreased from pluripotent cells to neurons. We identified 2400, 845, and 330 TIA1 mRNA targets in hESCs, NPC, and neurons, respectively. The vast majority of mRNA targets in hESC were genes associated with neurodevelopment and included autism spectrum disorder-risk genes that were not bound in neurons. Additionally, we found that most TIA1 mRNA targets have reduced ribosomal engagement levels. Conclusion Our results reveal TIA1 mRNA targets in hESCs and during human neurodevelopment, indicate that translation repression is a key process targeted by TIA1 binding and implicate TIA1 function in neuronal differentiation.

scholarly journals The Use of Peptides in the Treatment of Fragile X Syndrome: Challenges and Opportunities

2021 ◽  
Vol 12 ◽  
Author(s):  
Alice Romagnoli ◽  
Daniele Di Marino
Keyword(s):  
Fragile X Syndrome ◽  
Protein Interactions ◽  
Rna Binding ◽  
Fragile X ◽  
Pharmaceutical Research ◽  
Autism Spectrum ◽  
New Drugs ◽  
Protein Protein Interactions ◽  
Wide Range ◽  
Challenges And Opportunities

Fragile X Syndrome (FXS) is the most frequent cause of inherited intellectual disabilities and autism spectrum disorders, characterized by cognitive deficits and autistic behaviors. The silencing of the Fmr1 gene and consequent lack of FMRP protein, is the major contribution to FXS pathophysiology. FMRP is an RNA binding protein involved in the maturation and plasticity of synapses and its absence culminates in a range of morphological, synaptic and behavioral phenotypes. Currently, there are no approved medications for the treatment of FXS, with the approaches under study being fairly specific and unsatisfying in human trials. Here we propose peptides/peptidomimetics as candidates in the pharmacotherapy of FXS; in the last years this class of molecules has catalyzed the attention of pharmaceutical research, being highly selective and well-tolerated. Thanks to their ability to target protein-protein interactions (PPIs), they are already being tested for a wide range of diseases, including cancer, diabetes, inflammation, Alzheimer's disease, but this approach has never been applied to FXS. As FXS is at the forefront of efforts to develop new drugs and approaches, we discuss opportunities, challenges and potential issues of peptides/peptidomimetics in FXS drug design and development.

scholarly journals FMRP has a cell-type-specific role in CA1 pyramidal neurons to regulate autism-related transcripts and circadian memory

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Kirsty Sawicka ◽  
Caryn R Hale ◽  
Christopher Y Park ◽  
John J Fak ◽  
Jodi E Gresack ◽  
...  
Keyword(s):  
Pyramidal Neurons ◽  
Rna Binding ◽  
Fragile X ◽  
Neuronal Cell ◽  
Cell Types ◽  
Brain Regions ◽  
Cell Type ◽  
Ca1 Pyramidal Neurons ◽  
Mrna Targets ◽  
Specific Regulation

Loss of the RNA binding protein FMRP causes Fragile X Syndrome (FXS), the most common cause of inherited intellectual disability, yet it is unknown how FMRP function varies across brain regions and cell types and how this contributes to disease pathophysiology. Here we use conditional tagging of FMRP and CLIP (FMRP cTag CLIP) to examine FMRP mRNA targets in hippocampal CA1 pyramidal neurons, a critical cell type for learning and memory relevant to FXS phenotypes. Integrating these data with analysis of ribosome-bound transcripts in these neurons revealed CA1-enriched binding of autism-relevant mRNAs, and CA1-specific regulation of transcripts encoding circadian proteins. This contrasted with different targets in cerebellar granule neurons, and was consistent with circadian defects in hippocampus-dependent memory in Fmr1 knockout mice. These findings demonstrate differential FMRP-dependent regulation of mRNAs across neuronal cell types that may contribute to phenotypes such as memory defects and sleep disturbance associated with FXS.

scholarly journals Culture medium, gas atmosphere and MAPK inhibition affect regulation of RNA-binding protein targets during mouse preimplantation development

Reproduction ◽  
2011 ◽  
Vol 142 (5) ◽  
pp. 689-698 ◽  
Author(s):  
Michele D Calder ◽  
Patricia H Watson ◽  
Andrew J Watson
Keyword(s):  
Mrna Expression ◽  
Culture Medium ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Mapk Pathway ◽  
Preimplantation Development ◽  
Cell Stage ◽  
Target Mrnas ◽  
Mrna Targets ◽  
Gas Atmosphere

During oogenesis, mammalian oocytes accumulate maternal mRNAs that support the embryo until embryonic genome activation. RNA-binding proteins (RBP) may regulate the stability and turnover of maternal and embryonic mRNAs. We hypothesised that varying embryo culture conditions, such as culture medium, oxygen tension and MAPK inhibition, affects regulation of RBPs and their targets during preimplantation development. STAU1, ELAVL1, KHSRP and ZFP36 proteins and mRNAs were detected throughout mouse preimplantation development, whereasElavl2mRNA decreased after the two-cell stage. Potential target mRNAs of RBP regulation,Gclc,Slc2a1andSlc7a1were detected during mouse preimplantation development.GclcmRNA was significantly elevated in embryos cultured in Whitten's medium compared with embryos cultured in KSOMaa, andGclcmRNA was elevated under high-oxygen conditions. Inhibition of the p38 MAPK pathway reducedSlc7a1mRNA expression while inhibition of ERK increasedSlc2a1mRNA expression. The half-lives of the potential RBP mRNA targets are not regulated in parallel;Slc2a1mRNA displayed the longest half-life. Our results indicate that mRNAs and proteins encoding five RBPs are present during preimplantation development and more importantly, demonstrate that expression of RBP target mRNAs are regulated by culture medium, gas atmosphere and MAPK pathways.

scholarly journals A new strategy to uncover fragile X proteomic biomarkers using the nascent proteome of peripheral blood mononuclear cells (PBMCs)

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Olivier Dionne ◽  
François Corbin
Keyword(s):  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Rna Binding ◽  
Mononuclear Cells ◽  
Fragile X ◽  
Enrichment Analysis ◽  
Autism Spectrum ◽  
Label Free ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

AbstractFragile X syndrome (FXS) is the most prevalent inherited cause of intellectual disabilities and autism spectrum disorders. FXS result from the loss of expression of the FMRP protein, an RNA-binding protein that regulates the expression of key synaptic effectors. FXS is also characterized by a wide array of behavioural, cognitive and metabolic impairments. The severity and penetrance of those comorbidities are extremely variable, meaning that a considerable phenotypic heterogeneity is found among fragile X individuals. Unfortunately, clinicians currently have no tools at their disposal to assay a patient prognosis upon diagnosis. Since the absence of FMRP was repeatedly associated with an aberrant protein synthesis, we decided to study the nascent proteome in order to screen for potential proteomic biomarkers of FXS. We used a BONCAT (Biorthogonal Non-canonical Amino Acids Tagging) method coupled to label-free mass spectrometry to purify and quantify nascent proteins of peripheral blood mononuclear cells (PBMCs) from 7 fragile X male patients and 7 age-matched controls. The proteomic analysis identified several proteins which were either up or downregulated in PBMCs from FXS individuals. Eleven of those proteins were considered as potential biomarkers, of which 5 were further validated by Western blot. The gene ontology enrichment analysis highlighted molecular pathways that may contribute to FXS physiopathology. Our results suggest that the nascent proteome of PBMCs is well suited for the discovery of FXS biomarkers.

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