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Osterweil, E. K.

Publications and source records attributed to Osterweil, E. K..

7 recordsLinked to original sources

The most common epilepsy-causing mutation in EEF1A2 (E122K) perturbs the translation of specific transcripts but not the rate of global protein synthesis

Heterozygous de novo missense mutations in the EEF1A2 gene encoding translation elongation factor eEF1A2 result in neurodevelopmental disorders, typically characterised by early onset epilepsy and intellectual disability (ID). The E122K mutation is the most commonly reported missense mutation and is amongst the more severe in terms of epilepsy and ID. Here we made use of a recently developed mouse model which recapitulates the E122K mutation to examine how mutations in EEF1A2 might disrupt neuronal gene expression. Primary neurons from mutant mice and transfected HEK293T cells were used to examine effects on global protein synthesis. In contrast to previous reports, we were unable to detect a change in global protein synthesis using either of two different assay systems. TRAP-seq and mass spectrometry were then employed to study the effects of the mutation on the translatome and proteome respectively. These analyses revealed perturbation of expression of a subset of genes, with a slight skew towards downregulation, particularly for longer transcripts. Further analysis indicated a down regulation of proteins involved in synaptic function in both the translatomic and proteomic datasets.

neuroscience↗

FMR1 gene therapy restores activity-driven inhibition and prevents audiogenic seizures in Fmr1-/y mice

Fragile X syndrome (FXS) is a neurodevelopmental disorder associated with auditory hypersensitivity, circuit hyperexcitability, and seizures. Whether re-expression of the FMR1 gene and encoded Fragile X Messenger Ribonucleoprotein (FMRP) can restore sensory circuit dysfunction remains unclear. Here, we show that a viral AAV-FMR1 vector rescues audiogenic seizures in the Fmr1-/y mouse model after both neonatal and adult delivery, indicating that auditory circuit dysfunction remains reversible. Local re-expression in the inferior colliculus (IC) is sufficient to reduce seizure susceptibility, identifying this region as a key site of FMRP-dependent circuit regulation. In the IC, Translating Ribosome Affinity Purification and RNA-seq (TRAP-seq) profiling reveals impaired induction of sound-evoked translation programs in Fmr1-/y neurons, including those regulated by transcription factor Npas4. AAV-FMR1 restores a WT-like molecular response and normalizes unbalanced sound-evoked activation of VGLUT2+ excitatory neurons over VGAT+ inhibitory neurons in Fmr1-/y IC. Together, these findings indicate altered translation of Npas4 in response to sound impairs recruitment of inhibition in the Fmr1-/y IC, and this can be reversed with AAV-FMR1 administration. Moreover, the rescue of seizures after adult administration of AAV-FMR1 supports a gene therapy approach for FXS. HighlightsO_LITRAP-seq reveals impaired activity-driven translation in Fmr1-/y inferior colliculus (IC) C_LIO_LIDeficient Npas4 induction reduces evoked inhibition in Fmr1-/y IC C_LIO_LIAAV9-FMR1 gene therapy normalizes translation and excitatory/inhibitory balance in Fmr1-/y IC C_LIO_LIAAV9-FMR1 gene therapy prevents audiogenic seizures in Fmr1-/y mice when administered neonatally or in adulthood C_LI

neuroscience↗

FMR1 reduction alters cellular and circuit properties in human cortex

Transcriptional silencing of FMR1 results in Fragile X syndrome (FXS), the leading inherited cause of intellectual disability (ID) and autism. The Fmr1-/y mouse model has been used to identify FXS disease mechanisms, whereas mechanistic insights from human brain are lacking. By leveraging organotypic human cortical slices and viral tools to reduce FMR1 expression, we create a new model that captures cell type-specific transcriptomic changes similar to FXS patient cortex that are not seen in the Fmr1-/y mouse. Among these are ion channel subunit changes in deep layer pyramidal neurons, which are consistent with a robust hyperexcitability seen by whole-cell patch-clamp recordings, and increased synchronized activity revealed by 2-photon calcium imaging. Together, this work defines the impact of FMR1 reduction in human cortex and provides a new model for testing therapeutic interventions in FXS.

neuroscience↗

Syngap+/- CA1 pyramidal neurons exhibit upregulated translation of long mRNAs associated with LTP

In the Syngap+/- model of SYNGAP1-related intellectual disability (SRID), excessive neuronal protein synthesis is linked to deficits in synaptic plasticity. Here, we use Translating Ribosome Affinity Purification and RNA-seq (TRAP-seq) to identify mistranslating mRNAs in Syngap+/- CA1 pyramidal neurons that exhibit enhanced synaptic stability and impaired long-term potentiation (LTP). We find the translation environment is significantly altered in a manner that is distinct from the Fmr1-/y model of Fragile X Syndrome (FXS), another monogenic model of autism and intellectual disability (ID). The Syngap+/- translatome is enriched for regulators of DNA repair, and mimics changes induced with chemical LTP (cLTP) in WT. This includes a striking upregulation in the translation of mRNAs with a longer length (>2kb) coding sequence (CDS). In contrast, long CDS transcripts are downregulated with induction of Gp1 metabotropic glutamate receptor induced long-term depression (mGluR-LTD) in WT, and this profile is mimicked in the Fmr1-/y model. Together, our results show the Syngap+/- and Fmr1-/y models mimic the translation environments of LTP and LTD, respectively, consistent with the dysregulation of these plasticity states in each model. Moreover, we show that translation of >2kb mRNAs is a defining feature of LTP that is oppositely regulated during LTD, revealing a novel mRNA signature of plasticity.

neuroscience↗

Non-ionotropic signaling through the NMDA receptor GluN2B carboxy terminal domain drives morphological plasticity of dendritic spines and reverses fragile X phenotypes in mouse hippocampus

It is well known that activation of NMDA receptors can trigger long-term synaptic depression (LTD) and that a morphological correlate of this functional plasticity is spine retraction and elimination. Recent studies have led to the surprising conclusion that NMDA-induced spine shrinkage proceeds independently of ion flux and requires the initiation of de novo protein synthesis, highlighting an unappreciated contribution of mRNA translation to non-ionotropic NMDAR signaling. Here we used NMDA-induced spine shrinkage in slices of mouse hippocampus as a readout to investigate this novel modality of synaptic transmission. By using selective pharmacological and genetic tools, we find that structural plasticity is dependent on the ligand binding domain (LBD) of GluN2B-containing NMDA receptors and that metabotropic signaling occurs via the GluN2B carboxyterminal domain (CTD). Disruption of signaling by replacing the GluN2B CTD with the GluN2A CTD leads to increased spine density, dysregulated basal protein synthesis, and epileptiform activity in area CA3 reminiscent of phenotypes observed in the Fmr1-/y model of fragile X syndrome. By crossing the Fmr1-/y mice with animals in which the GluN2A CTD has been replaced with the GluN2B CTD, we observe a correction of these core fragile X phenotypes. These findings suggest that non-ionotropic NMDAR signaling through GluN2B may represent a novel therapeutic target for treatment of fragile X and related causes of intellectual disability and autism.

neuroscience↗

Excess ribosomal protein production unbalances translation in Fragile X Syndrome

Dysregulated protein synthesis is a core pathogenic mechanism in Fragile X Syndrome (FX). The mGluR Theory of FX predicts that pathological synaptic changes arise from the excessive translation of mRNAs downstream of mGlu1/5 activation. Here, we use a combination of CA1 pyramidal neuron-specific TRAP-seq and proteomics to identify the overtranslating mRNAs supporting exaggerated mGlu1/5-induced long-term synaptic depression (mGluR-LTD) in the FX mouse model (Fmr1-/y). Surprisingly, our results identify a robust translation of ribosomal proteins (RPs) upon mGlu1/5 stimulation that coincides with a reduced translation of long mRNAs encoding synaptic proteins. These changes are mimicked and occluded in Fmr1-/y neurons. Inhibiting RP translation significantly impairs mGluR-LTD and prevents the length-dependent shift in the translating population. Together, these results suggest that pathological changes in FX result from a length-dependent alteration in the translating population that is supported by excessive RP translation.

neuroscience↗

Lovastatin, not simvastatin, corrects core phenotypes in the fragile X mouse model

The cholesterol-lowering drug lovastatin corrects neurological phenotypes in animal models of fragile X syndrome (FX), a commonly identified genetic cause of autism and intellectual disability. The therapeutic efficacy of lovastatin is being tested in clinical trials for FX, however the structurally similar drug simvastatin has been proposed as an alternative due to an increased potency and brain penetrance. Here, we perform a side-by-side comparison of the effects of lovastatin and simvastatin treatment on two core phenotypes in the Fmr1-/y mouse model. We find that while lovastatin normalizes excessive hippocampal protein synthesis and reduces audiogenic seizures (AGS) in the Fmr1-/y mouse, simvastatin does not correct either phenotype. These results caution against the assumption that simvastatin is a valid alternative to lovastatin for the treatment of FX.

neuroscience↗