Argonaute: A Voyage into the Role of Protein
Phosphorylation in Synaptic Plasticity and Memory

BACKGROUND:

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Argonaute (Ago2) is not only a protein with a pretty neat name (this name being derived from a funky mutation in Arabidopsis rather than the Greek myth!), but also seems to contribute to normal brain function. Argonaute is a core member of the RNA-induced silencing complex, or RISC, which is a group of proteins that work together to prevent specific messenger RNA transcripts from encountering a ribosome in the cytoplasm and being translated. Work from our lab has previously shown that Argonaute is phosphorylated following the induction of long-term depression, which appears to block the translation of mRNA transcripts belonging to a protein normally involved in regulating the actin cytoskeleton of dendritic spines. This leads to spine shrinkage, which is a key hallmark of this form of synaptic plasticity. Since these experiments were largely done in vitro, the purpose of my research is to fundamentally identify if these findings are conserved in vivo using a novel transgenic mouse model, in which the serine-387 residue in Argonaute phosphorylated during LTD has been mutated to the non-phosphorylatable residue alanine (the ‘Ago2 S388A’ strain). I am additionally investigating how this contributes to different forms of memory dependent on appropriate long-term depression in the hippocampus and perirhinal cortex of these mice, and am attempting to take a closer look at what is happening within the dendritic spines of neurons in these brain regions at a molecular level.

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METHODOLOGY:

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Since I’m primarily based in biochemistry, I’ve so far carried out *many* western blots and co-immunoprecipitations to quantify protein expression and protein:protein interactions in whole tissue and synaptosomal lysates from my mice. As I’m expecting to see reduced spine shrinkage following long-term depression in the Ago2 S388A mice, I’ve also spent some time refining and utilising the Golgi-cox staining technique. This (VERY affordable!) technique randomly stains a small percentage of neurons in a brain sample black, allowing you to pick out and measure structural features quite easily (such as spines). I’ve also carried out a range of object-based exploration assays and maze-based tasks to determine how preventing argonaute phosphorylation affects memory function. Right now, I’m based in one of my co-supervisor's electrophysiology labs in Exeter measuring field EPSPs to explore a hypothesised deficit in LTD.

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RESULTS:

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So far, my research has yielded two main findings. The first is that I’ve been able to confirm an upregulation in the expression of the protein involved in regulating the actin cytoskeleton of spines previously shown to be targeted by RISC-mediated pre-translational repression following Ago2 S387 phosphorylation in vitro, in both the hippocampus and cortex of Ago2 S388A mice. The second is that these mice show deficits in short-term spatial memory, suggesting a largely hippocampal-dependent role for Ago2 S387 phosphorylation.

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