The integrated stress response effector GADD34 is repurposed by neurons to promote stimulus-induced translation.

Neuronal protein synthesis is required for long-lasting plasticity and long-term memory consolidation. Dephosphorylation of eukaryotic initiation factor 2α is one of the key translational control events that is required to increase de novo protein synthesis that underlies long-lasting plasticity and memory consolidation. Here, we interrogate the molecular pathways of translational control that are triggered by neuronal stimulation with brain-derived neurotrophic factor (BDNF), which results in eukaryotic initiation factor 2α (eIF2α) dephosphorylation and increases in de novo protein synthesis. Primary rodent neurons exposed to BDNF display elevated translation of GADD34, which facilitates eIF2α dephosphorylation and subsequent de novo protein synthesis. Furthermore, GADD34 requires G-actin generated by cofilin to dephosphorylate eIF2α and enhance protein synthesis. Finally, GADD34 is required for BDNF-induced translation of synaptic plasticity-related proteins. Overall, we provide evidence that neurons repurpose GADD34, an effector of the integrated stress response, as an orchestrator of rapid increases in eIF2-dependent translation in response to plasticity-inducing stimuli.

Age-dependent shift in the de novo proteome accompanies pathogenesis in an Alzheimer's disease mouse model.

Alzheimer's disease (AD) is an age-related neurodegenerative disorder associated with memory loss, but the AD-associated neuropathological changes begin years before memory impairments. Investigation of the early molecular abnormalities in AD might offer innovative opportunities to target memory impairment prior to onset. Decreased protein synthesis plays a fundamental role in AD, yet the consequences of this dysregulation for cellular function remain unknown. We hypothesize that alterations in the de novo proteome drive early metabolic alterations in the hippocampus that persist throughout AD progression. Using a combinatorial amino acid tagging approach to selectively label and enrich newly synthesized proteins, we found that the de novo proteome is disturbed in young APP/PS1 mice prior to symptom onset, affecting the synthesis of multiple components of the synaptic, lysosomal, and mitochondrial pathways. Furthermore, the synthesis of large clusters of ribosomal subunits were affected throughout development. Our data suggest that large-scale changes in protein synthesis could underlie cellular dysfunction in AD.

Secreted Amyloid Precursor Protein-Alpha Enhances LTP Through the Synthesis and Trafficking of Ca2+-Permeable AMPA Receptors.

Regulation of AMPA receptor expression by neuronal activity and neuromodulators is critical to the expression of both long-term potentiation (LTP) and memory. In particular, Ca2+-permeable AMPARs (CP-AMPAR) play a unique role in these processes due to their transient, activity-regulated expression at synapses. Secreted amyloid precursor protein-alpha (sAPPα), a metabolite of the parent amyloid precursor protein (APP) has been previously shown to enhance hippocampal LTP as well as memory formation in both normal animals and in Alzheimer's disease models. In earlier work we showed that sAPPα promotes trafficking of GluA1-containing AMPARs to the cell surface and specifically enhances synthesis of GluA1. To date it is not known whether de novo synthesized GluA1 form CP-AMPARs or how they contribute to sAPPα-mediated plasticity. Here, using fluorescent non-canonical amino acid tagging-proximity ligation assay (FUNCAT-PLA), we show that brief treatment of primary rat hippocampal neurons with sAPPα (1 nM, 30 min) rapidly enhanced the cell-surface expression of de novo GluA1 homomers and reduced levels of de novo GluA2, as well as extant GluA2/3-AMPARs. The de novo GluA1-containing AMPARs were localized to extrasynaptic sites and later internalized by sAPPα-driven expression of the activity-regulated cytoskeletal-associated protein, Arc. Interestingly, longer exposure to sAPPα increased synaptic levels of GluA1/2 AMPARs. Moreover, the sAPPα-mediated enhancement of LTP in area CA1 of acute hippocampal slices was dependent on CP-AMPARs. Together, these findings show that sAPPα engages mechanisms which specifically enhance the synthesis and cell-surface expression of GluA1 homomers, underpinning the sAPPα-driven enhancement of synaptic plasticity in the hippocampus.

Docosahexaenoic acid prevents palmitate-induced insulin-dependent impairments of neuronal health.

The importance of fatty acids (FAs) for healthy brain development and function has become more evident in the past decades. However, most studies focus on the hypothalamus as an important FA-sensing brain region involved in energy homeostasis. Less work has been done to evaluate the effects of FAs on brain regions such as the hippocampus or cortex, two important centres of learning, memory formation, and cognition. Furthermore, the mechanisms of how FAs modulate the neuronal development and function are incompletely understood. Therefore, this study examined the effects of the saturated FA palmitic acid (PA) and the polyunsaturated FA docosahexaenoic acid (DHA) on primary hippocampal and cortical cultures isolated from P0/P1 Sprague Dawley rat pups. Exposure to PA, but not DHA, resulted in severe morphological changes in primary neurons such as cell body swelling, axonal and dendritic blebbing, and a reduction in synaptic innervation, compromising healthy cell function and excitability. Pharmacological assessment revealed that the PA-mediated alterations were caused by overactivation of neuronal insulin signaling, demonstrated by insulin stimulation and phosphoinositide 3-kinase inhibition. Remarkably, co-exposure to DHA prevented all PA-induced morphological changes. This work provides new insights into how FAs can affect the cytoskeletal rearrangements and neuronal function via modulation of insulin signaling.

Suppression of MEHMO Syndrome Mutation in eIF2 by Small Molecule ISRIB.

Dysregulation of cellular protein synthesis is linked to a variety of diseases. Mutations in EIF2S3, encoding the γ subunit of the heterotrimeric eukaryotic translation initiation factor eIF2, cause MEHMO syndrome, an X-linked intellectual disability disorder. Here, using patient-derived induced pluripotent stem cells, we show that a mutation at the C terminus of eIF2γ impairs CDC123 promotion of eIF2 complex formation and decreases the level of eIF2-GTP-Met-tRNAiMet ternary complexes. This reduction in eIF2 activity results in dysregulation of global and gene-specific protein synthesis and enhances cell death upon stress induction. Addition of the drug ISRIB, an activator of the eIF2 guanine nucleotide exchange factor, rescues the cell growth, translation, and neuronal differentiation defects associated with the EIF2S3 mutation, offering the possibility of therapeutic intervention for MEHMO syndrome.

Secreted Amyloid Precursor Protein-Alpha Promotes Arc Protein Synthesis in Hippocampal Neurons.

Secreted amyloid precursor protein-α (sAPPα) is a neuroprotective and memory-enhancing molecule, however, the mechanisms through which sAPPα promotes these effects are not well understood. Recently, we have shown that sAPPα enhances cell-surface expression of glutamate receptors. Activity-related cytoskeletal-associated protein Arc (Arg3.1) is an immediate early gene capable of modulating long-term potentiation, long-term depression and homeostatic plasticity through regulation of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor localization. Accordingly, we hypothesized that sAPPα may enhance synaptic plasticity, in part, by the de novo synthesis of Arc. Using primary cortical and hippocampal neuronal cultures we found that sAPPα (1 nM, 2 h) enhances levels of Arc mRNA and protein. Arc protein levels were increased in both the neuronal somata and dendrites in a Ca2+/calmodulin-dependent protein kinase II-dependent manner. Additionally, dendritic Arc expression was dependent upon activation of mitogen-activated protein kinase and protein kinase G. The enhancement of dendritic Arc protein was significantly reduced by antagonism of N-methyl-D-aspartate (NMDA) and nicotinic acetylcholine (α7nACh) receptors, and fully eliminated by dual application of these antagonists. This effect was further corroborated in area CA1 of acute hippocampal slices. These data suggest sAPPα-regulated plasticity within hippocampal neurons is mediated by cooperation of NMDA and α7nACh receptors to engage a cascade of signal transduction molecules to enhance the transcription and translation of Arc.

Glutamate Receptor Trafficking and Protein Synthesis Mediate the Facilitation of LTP by Secreted Amyloid Precursor Protein-Alpha.

Secreted amyloid precursor protein-alpha (sAPPα) has growth factor-like properties and can modulate long-term potentiation (LTP) and memory. Here, we demonstrate that exposure to sAPPα converts short-lasting LTP into protein-synthesis-dependent late LTP in hippocampal slices from male rats. sAPPß had no discernable effect. We hypothesized that sAPPα facilitated LTP via regulated glutamate receptor trafficking and de novo protein synthesis. We found using a linear mixed model that sAPPα stimulated trafficking of GluA2-lacking AMPARs, as well as NMDARs to the extrasynaptic cell surface, in a calcium/calmodulin-dependent kinase II and protein kinase G-dependent manner. Both cell surface receptor accumulation and LTP facilitation were present even after sAPPα washout and inhibition of receptor trafficking or protein synthesis prevented all these effects. Direct visualization of newly synthesized proteins (FUNCAT-PLA) confirmed the ability of sAPPα to stimulate de novo protein synthesis and revealed GluA1 as one of the upregulated proteins. Therefore, sAPPα generates a coordinated synthesis and trafficking of glutamate receptors to the cell surface that facilitate LTP.SIGNIFICANCE STATEMENT Secreted amyloid precursor protein-alpha (sAPPα) is a neurotrophic and neuroprotective protein that can promote synaptic plasticity and memory, yet the molecular mechanisms underlying these effects are still not well understood. Here, we show that sAPPα facilitates long-term potentiation (LTP) in a concentration-dependent fashion through cellular processes involving de novo protein synthesis and trafficking of both GluA2-lacking AMPARs and NMDARs to the extrasynaptic cell surface. sAPPα also enhances GluA1, but not GluA2, synthesis. The trafficking effects, along with the LTP facilitation, persist after sAPPα washout, revealing a metaplastic capability of exogenous sAPPα administration. sAPPα thus facilitates LTP through coordinated activation of protein synthesis and trafficking of glutamate receptors to the cell surface, where they are positioned for priming LTP.