strategies-for-assessing-autistic-like-behaviors-in-mice

Strategies for Assessing Autistic-Like Behaviors in Mice

Autism spectrum disorder (ASD) is a neurobiologically complex condition with a heterogeneous genetic etiology. Clinically, ASD is diagnosed by social ...

McGill University

Fragile X syndrome (FXS) is the leading monogenic cause of autism spectrum disorders (ASD). Trinucleotide repeat expansions in FMR1 abolish FMRP expression, leading to hyperactivation of ERK and mTOR signaling upstream of mRNA translation. Here we show that metformin, the most widely used drug for type 2 diabetes, rescues core phenotypes in Fmr1 mice and selectively normalizes ERK signaling, eIF4E phosphorylation and the expression of MMP-9. Thus, metformin is a potential FXS therapeutic.

Metformin ameliorates core deficits in a mouse model of fragile X syndrome.

Translation of mRNA into protein has a fundamental role in neurodevelopment, plasticity, and memory formation; however, its contribution in the pathophysiology of depressive disorders is not fully understood. We investigated the involvement of MNK1/2 (MAPK-interacting serine/threonine-protein kinase 1 and 2) and their target, eIF4E (eukaryotic initiation factor 4E), in depression-like behavior in mice. Mice carrying a mutation in eIF4E for the MNK1/2 phosphorylation site (Ser209Ala, Eif4e ki/ki), the Mnk1/2 double knockout mice (Mnk1/2), or mice treated with the MNK1/2 inhibitor, cercosporamide, displayed anxiety- and depression-like behaviors, impaired serotonin-induced excitatory synaptic activity in the prefrontal cortex, and diminished firing of the dorsal raphe neurons. In Eif4e ki/ki mice, brain IκBα, was decreased, while the NF-κB target, TNFα was elevated. TNFα inhibition in Eif4e ki/ki mice rescued, whereas TNFα administration to wild-type mice mimicked the depression-like behaviors and 5-HT synaptic deficits. We conclude that eIF4E phosphorylation modulates depression-like behavior through regulation of inflammatory responses.

Translational control of depression-like behavior via phosphorylation of eukaryotic translation initiation factor 4E.

Translational control plays a key role in regulation of neuronal activity and behavior. Deletion of the translational repressor 4E-BP2 in mice alters excitatory and inhibitory synaptic functions, engendering autistic-like behaviors. The contribution of 4E-BP2-dependent translational control in excitatory and inhibitory neurons and astrocytic cells to these behaviors remains unknown. To investigate this, we generated cell-type-specific conditional 4E-BP2 knockout mice and tested them for the salient features of autism, including repetitive stereotyped behaviors (self-grooming and marble burying), sociability (3-chamber social and direct social interaction tests), and communication (ultrasonic vocalizations in pups). We found that deletion of 4E-BP2 in GABAergic inhibitory neurons, defined by , resulted in impairments in social interaction and vocal communication. In contrast, deletion of 4E-BP2 in forebrain glutamatergic excitatory neurons, defined by , or in astrocytes, defined by , failed to cause autistic-like behavioral abnormalities. Taken together, we provide evidence for an inhibitory-cell-specific role of 4E-BP2 in engendering autism-related behaviors.

Inhibitory interneurons mediate autism-associated behaviors via 4E-BP2.

Control of protein synthesis (mRNA translation) is essential for proper brain development and function. Perturbations to the mechanisms governing mRNA translation have repeatedly been shown to constitute a neurodegenerative, neuropsychiatric, and neurodevelopmental disorder risk factor. Developing effective therapeutics for brain disorders will require a better understanding of the molecular mechanisms underlying the control of protein synthesis in brain function. Studies using transgenic animal models have been invaluable towards this end, providing exciting new insights into the genetic basis of brain disorders with hopeful prospects for new and effective treatment options.

Dysregulated translational control in brain disorders: from genes to behavior.

The regulation of protein synthesis is a critical step in gene expression, and its dysfunction is implicated in autism spectrum disorder (ASD). The eIF4E homologous protein (4EHP, also termed eIF4E2) binds to the mRNA 5' cap to repress translation. The stability of 4EHP is maintained through physical interaction with GRB10 interacting GYF protein 2 (GIGYF2). Gene-disruptive mutations in GIGYF2 are linked to ASD, but causality is lacking. We hypothesized that GIGYF2 mutations cause ASD by disrupting 4EHP function.

The eIF4E homolog 4EHP (eIF4E2) regulates hippocampal long-term depression and impacts social behavior.

Type I interferons (IFNs) are critical cytokines in the host defense against invading pathogens. Sustained production of IFNs, however, is detrimental to the host, as it provokes autoimmune diseases. Thus, the expression of IFNs is tightly controlled. We report that the mRNA 5' cap-binding protein 4EHP plays a key role in regulating type I IFN concomitant with controlling virus replication, both in vitro and in vivo. Mechanistically, 4EHP suppresses IFN-β production by effecting the miR-34a-induced translational silencing of Ifnb1 mRNA. miR-34a is upregulated by both RNA virus infection and IFN-β induction, prompting a negative feedback regulatory mechanism that represses IFN-β expression via 4EHP. These findings demonstrate the direct involvement of 4EHP in virus-induced host response, underscoring a critical translational silencing mechanism mediated by 4EHP and miR-34a to impede sustained IFN production. This study highlights an intrinsic regulatory function for miRNA and the translation machinery in maintaining host homeostasis.

microRNA-induced translational control of antiviral immunity by the cap-binding protein 4EHP.

Viruses evade the innate immune response by suppressing the production or activity of cytokines such as type I interferons (IFNs). Here we report the discovery of a mechanism by which the SARS-CoV-2 virus coopts an intrinsic cellular machinery to suppress the production of the key immunostimulatory cytokine IFN-β. We reveal that the SARS-CoV-2 encoded nonstructural protein 2 (NSP2) directly interacts with the cellular GIGYF2 protein. This interaction enhances the binding of GIGYF2 to the mRNA cap-binding protein 4EHP, thereby repressing the translation of the  mRNA. Depletion of GIGYF2 or 4EHP significantly enhances IFN-β production, which inhibits SARS-CoV-2 replication. Our findings reveal a target for rescuing the antiviral innate immune response to SARS-CoV-2 and other RNA viruses.

SARS-CoV-2 impairs interferon production via NSP2-induced repression of mRNA translation.

The consolidation of learned information into long-lasting memories requires the strengthening of synaptic connections through de novo protein synthesis. Translation initiation factors play a cardinal role in gating the production of new proteins thereby regulating memory formation. Both positive and negative regulators of translation play a critical role in learning and memory consolidation. The eukaryotic initiation factor 4E (eIF4E) homologous protein (4EHP, encoded by the gene Eif4e2) is a pivotal negative regulator of translation but its role in learning and memory is unknown. To address this gap in knowledge, we generated excitatory (glutamatergic: CaMKIIα-positive) and inhibitory (GABAergic: GAD65-positive) conditional knockout mice for 4EHP, which were analyzed in various behavioral memory tasks. Knockout of 4EHP in Camk2a-expressing neurons (4EHP-cKO) did not impact long-term memory in either contextual fear conditioning or Morris water maze tasks. Similarly, long-term contextual fear memory was not altered in Gad2-directed 4EHP knockout mice (4EHP-cKO). However, when subjected to a short-term T-maze working memory task, both mouse models exhibited impaired cognition. We therefore tested the hypothesis that de novo protein synthesis plays a direct role in working memory. We discovered that phosphorylation of ribosomal protein S6, a measure of mTORC1 activity, is dramatically reduced in the CA1 hippocampus of 4EHP-cKO mice. Consistently, genetic reduction of mTORC1 activity in either excitatory or inhibitory neurons was sufficient to impair working memory. Taken together, these findings indicate that translational control by 4EHP and mTORC1 in both excitatory and inhibitory neurons are necessary for working memory.

Cell-type-specific translational control of spatial working memory by the cap-binding protein 4EHP.

Impaired brain protein synthesis, synaptic plasticity, and memory are major hallmarks of Alzheimer's disease (AD). The ketamine metabolite (2R,6R)-hydroxynorketamine (HNK) has been shown to modulate protein synthesis, but its effects on memory in AD models remain elusive.

The ketamine metabolite (2R,6R)-hydroxynorketamine rescues hippocampal mRNA translation, synaptic plasticity and memory in mouse models of Alzheimer's disease.

Article	Year
Metformin ameliorates core deficits in a mouse model of fragile X syndrome. Nature medicine\| PubMed ID: 28504725	2017
Translational control of depression-like behavior via phosphorylation of eukaryotic translation initiation factor 4E. Nature communications\| PubMed ID: 29941989	2018
Inhibitory interneurons mediate autism-associated behaviors via 4E-BP2. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 31427534	2019
Dysregulated translational control in brain disorders: from genes to behavior. Current opinion in genetics & development\| PubMed ID: 32535350	2020
The eIF4E homolog 4EHP (eIF4E2) regulates hippocampal long-term depression and impacts social behavior. Molecular autism\| PubMed ID: 33225984	2020
microRNA-induced translational control of antiviral immunity by the cap-binding protein 4EHP. Molecular cell\| PubMed ID: 33581076	2021
SARS-CoV-2 impairs interferon production via NSP2-induced repression of mRNA translation. Proceedings of the National Academy of Sciences of the United States of America\| PubMed ID: 35878012	2022
Cell-type-specific translational control of spatial working memory by the cap-binding protein 4EHP. Molecular brain\| PubMed ID: 36650535	2023
The ketamine metabolite (2R,6R)-hydroxynorketamine rescues hippocampal mRNA translation, synaptic plasticity and memory in mouse models of Alzheimer's disease. Alzheimer's & dementia : the journal of the Alzheimer's Association\| PubMed ID: 38934107	2024

Shane Wiebe

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