Differential regulation of local mRNA dynamics and translation following long-term potentiation and depression

Decades of work have demonstrated that messenger RNAs (mRNAs) are localized and translated within neuronal dendrites and axons to provide proteins for remodeling and maintaining growth cones or synapses. It remains unknown, however, whether specific forms of plasticity differentially regulate the dynamics and translation of individual mRNA species. To address this, we targeted three individual synaptically localized mRNAs, CamkIIa, β-actin, Psd95, and used molecular beacons to track endogenous mRNA movements. We used reporters and CRISPR/Cas9 gene editing to track mRNA translation in cultured neurons. We found alterations in mRNA dynamic properties occurred during two forms of synaptic plasticity, long-term potentiation (cLTP) and depression (mGluR-LTD). Changes in mRNA dynamics following either form of plasticity resulted in an enrichment of mRNA in the vicinity of dendritic spines. Both the reporters and tagging of endogenous proteins revealed the transcript-specific stimulation of protein synthesis following cLTP or mGluR-LTD. As such, the plasticity-induced enrichment of mRNA near synapses could be uncoupled from its translational status. The enrichment of mRNA in the proximity of spines allows for localized signaling pathways to decode plasticity milieus and stimulate a specific translational profile, resulting in a customized remodeling of the synaptic proteome

The switch-like expression of heme-regulated kinase 1 mediates neuronal proteostasis following proteasome inhibition

We examined the feedback between the major protein degradation pathway, the ubiquitin-proteasome system (UPS), and protein synthesis in rat and mouse neurons. When protein degradation was inhibited, we observed a coordinate dramatic reduction in nascent protein synthesis in neuronal cell bodies and dendrites. The mechanism for translation inhibition involved the phosphorylation of eIF2alpha, surprisingly mediated by eIF2alpha kinase 1, or heme-regulated kinase inhibitor (HRI). Under basal conditions, neuronal expression of HRI is barely detectable. Following proteasome inhibition, HRI protein levels increase owing to stabilization of HRI and enhanced translation, likely via the increased availability of tRNAs for its rare codons. Once expressed, HRI is constitutively active in neurons because endogenous heme levels are so low; HRI activity results in eIF2alpha phosphorylation and the resulting inhibition of translation. These data demonstrate a novel role for neuronal HRI that senses and responds to compromised function of the proteasome to restore proteostasis

Overview of Current Drugs and Molecules in Development for Spinal Muscular Atrophy Therapy

The full text of this article can be found here: https://link.springer.com/article/10.1007/s40265-018-0868-

Local translation in neuronal processes

Neurons exhibit a unique degree of spatial compartmentalization and are able to maintain and remodel their proteomes independently from the cell body. While much effort has been devoted to understanding the capacity and role for local protein synthesis in dendrites and spines, local mRNA translation in mature axons, projecting over distances up to a meter, has received much less attention. Also, little is known about the spatio-temporal dynamics of axonal and dendritic gene expression as function of mRNA abundance, protein synthesis and degradation. Here, we summarize key recent findings that have shaped our knowledge of the precise location of local protein production and discuss unique strategies used by neurons to shape presynaptic and postsynaptic proteomes

Dynamics of survival of motor neuron (SMN) protein interaction with the mRNA-binding protein IMP1 facilitates its trafficking into motor neuron axons

Spinal muscular atrophy (SMA) is a lethal neurodegenerative disease specifically affecting spinal motor neurons. SMA is caused by the homozygous deletion or mutation of the survival of motor neuron 1 (SMN1) gene. The SMN protein plays an essential role in the assembly of spliceosomal ribonucleoproteins. However, it is still unclear how low levels of the ubiquitously expressed SMN protein lead to the selective degeneration of motor neurons. An additional role for SMN in the regulation of the axonal transport of mRNA-binding proteins (mRBPs) and their target mRNAs has been proposed. Indeed, several mRBPs have been shown to interact with SMN, and the axonal levels of few mRNAs, such as the beta-actin mRNA, are reduced in SMA motor neurons. In this study we have identified the beta-actin mRNA-binding protein IMP1/ZBP1 as a novel SMN-interacting protein. Using a combination of biochemical assays and quantitative imaging techniques in primary motor neurons, we show that IMP1 associates with SMN in individual granules that are actively transported in motor neuron axons. Furthermore, we demonstrate that IMP1 axonal localization depends on SMN levels, and that SMN deficiency in SMA motor neurons leads to a dramatic reduction of IMP1 protein levels. In contrast, no difference in IMP1 protein levels was detected in whole brain lysates from SMA mice, further suggesting neuron specific roles of SMN in IMP1 expression and localization. Taken together, our data support a role for SMN in the regulation of mRNA localization and axonal transport through its interaction with mRBPs such as IMP1

Dynamics of survival of motor neuron (SMN) protein interaction with the mRNA-binding protein IMP1 facilitates its trafficking into motor neuron axons

Spinal muscular atrophy (SMA) is a lethal neurodegenerative disease specifically affecting spinal motor neurons. SMA is caused by the homozygous deletion or mutation of the survival of motor neuron 1 (SMN1) gene. The SMN protein plays an essential role in the assembly of spliceosomal ribonucleoproteins. However, it is still unclear how low levels of the ubiquitously expressed SMN protein lead to the selective degeneration of motor neurons. An additional role for SMN in the regulation of the axonal transport of mRNA-binding proteins (mRBPs) and their target mRNAs has been proposed. Indeed, several mRBPs have been shown to interact with SMN, and the axonal levels of few mRNAs, such as the beta-actin mRNA, are reduced in SMA motor neurons. In this study we have identified the beta-actin mRNA-binding protein IMP1/ZBP1 as a novel SMN-interacting protein. Using a combination of biochemical assays and quantitative imaging techniques in primary motor neurons, we show that IMP1 associates with SMN in individual granules that are actively transported in motor neuron axons. Furthermore, we demonstrate that IMP1 axonal localization depends on SMN levels, and that SMN deficiency in SMA motor neurons leads to a dramatic reduction of IMP1 protein levels. In contrast, no difference in IMP1 protein levels was detected in whole brain lysates from SMA mice, further suggesting neuron specific roles of SMN in IMP1 expression and localization. Taken together, our data support a role for SMN in the regulation of mRNA localization and axonal transport through its interaction with mRBPs such as IMP1

Correlating DNA-PAINT and single-molecule FRET for multiplexed super-resolution imaging

Correlating DNA-PAINT (point accumulation for imaging in nanoscale topography) and single-molecule FRET (Forster resonance energy transfer) enables the multiplexed detection with sub-diffraction optical resolution. We designed pairs of short oligonucleotides, labeled with donor and acceptor fluorophores with various distances generating different FRET efficiencies. The strands can transiently bind to a target docking strand, simultaneous binding of both strands results in FRET signals which yield a super-resolved image via DNA-PAINT imaging. We demonstrate FRET-PAINT by designing and imaging DNA origami, which is a useful tool to establish super-resolution methods. The DNA origami structures were equipped with three target binding sites spaced by 55 nm, a sub-diffraction limited distance, however ensuring that no FRET between the target sites occurs. We resolved the individual binding sites in the donor and acceptor channels, and in addition extracted the FRET efficiency for each site in single and mixed populations. The combination of FRET and DNA-PAINT allows for multiplexed super-resolution imaging in conjunction with distance-sensitive readout in the 1 to 10 nm range