MicroRNA control of local synaptic protein synthesis in neuronal dendrites

Lead Research Organisation: University of Bristol
Department Name: Biochemistry

Abstract

Neurons are a unique cell type because they are extremely polarised, with long processes extending millimetres from the nucleus in the cell body. This poses a cell biological challenge: how is protein expression at distal synapses regulated? One solution is that regulation of protein synthesis is decentralised, with local control of translation in dendrites to supply proteins according to the requirements of specific synapses.

MicroRNAs (miRNAs) are small, endogenous RNA molecules that repress the translation of target mRNAs by associating with Argonaute (Ago) proteins in the RNA-induced silencing complex (RISC) and are fundamentally important for fine-tuning protein synthesis in numerous cellular processes. Long-term synaptic plasticity underlies learning and memory by modifying neural circuitry, a major component of which is morphological changes of dendritic spines, which contain the postsynaptic machinery. Regulated miRNA activity plays a key role in this process by modulating the translation of cytoskeletal proteins that determine spine morphology. miRNA dysregulation is implicated in several neurological disorders that involve synaptic dysfunction, including Alzheimer's disease.

A key question is how "local" is this control of translation, i.e., does gene silencing spread along the dendrite to neighbouring unstimulated synapses, and if so, how is this regulated? This is important because dominant theories of Hebbian learning assume that plasticity is synapse-specific, while emerging evidence suggests otherwise. We have recently defined mechanisms for rapidly increasing miRNA-mediated gene silencing in response to NMDA receptor stimulation to cause dendritic spine shrinkage. Our hypothesis is that miRNA activity is modulated close to the stimulated spine to locally regulate translation and hence influence the morphology of only a small number of neighbouring spines.

The main experimental approach will be single-synapse stimulation by glutamate uncaging, followed by live (e.g. single-molecule imaging of nascent peptides using multimerized Sun-Tags) and fixed-cell (e.g. puromycin-proximity ligation assays) imaging techniques to analyse the translation of specific proteins and dynamic imaging of spine morphology in that region of dendrite. The project will investigate the mechanisms involved via mutagenesis of essential RISC proteins.
In addition, molecular-level computer simulation models will be developed to test our hypotheses and dissect parts of the system that are not experimentally dissociable. For example, separately comparing the roles of 1) diffusion of miRNA and signalling factors, 2) relative numbers of mRNAs, miRNAs and signalling factors,
3) dendrite and spine geometry for restricting protein translation to localised spatial compartments.

This joint experimental-computational project will define key mechanisms of local translation at synapses.

Publications

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Studentship Projects

Project Reference Relationship Related To Start End Student Name
BB/T008741/1 01/10/2020 30/09/2028
2429514 Studentship BB/T008741/1 21/09/2020 30/09/2024 Sofia Birgitta Raak