Award details

Structural, biochemical and computational studies of KIBRA protein-protein and protein-phospholipid interactions that are important for memory

Reference	BB/J008176/1
Principal Investigator / Supervisor	Dr Stefan Bagby
Co-Investigators / Co-Supervisors	Professor Jean van den Elsen
Institution	University of Bath
Department	Biology and Biochemistry
Funding type	Research
Value (£)	325,234
Status	Completed
Type	Research Grant
Start date	01/08/2012
End date	29/01/2016
Duration	42 months

Abstract

We will advance understanding of the molecular basis of KIBRA's role in memory as follows: Details underlying C2 domain role in KIBRA membrane trafficking For normal and variant KIBRA C2 domains, we will use NMR, isothermal titration calorimetry (ITC) and X-ray crystallography (XRC) to measure Ca2+ binding properties (affinity, stoichiometry, cooperativity, occupation order of Ca2+ sites (up to 3), and Ca2+-induced structural changes). Phospholipid specificity will be probed by NMR and ITC/surface plasmon resonance (SPR) measurement of normal and variant C2 binding to monodisperse analogues of phosphatidyl-inositol phosphates, -serine and -choline. We will use multiscale simulations to study membrane penetration depth, membrane-binding geometry, lipid determinants and role of different forces in normal and variant C2-membrane interaction. KIBRA WW domains We will determine whether the 2 tandem WW domains of KIBRA are cooperative or independent, ascertain the role of the atypical (Ile instead of Trp) second WW and establish the selectivity and structures of single and tandem WW interactions with single and multiple PPxY motif ligands from Dendrin and Synaptopodin, postsynaptic cytoskeleton organisers associated with neuronal synaptic plasticity. KIBRA-PKMzeta association PKMzeta is pivotal for long term memory storage. KIBRA-PKMzeta association is vital for PKMzeta function. We will use ITC, thermal shift, enzyme assays, XRC, NMR, and mutagenesis to delineate thermodynamics, kinetics, structural details and key residues of KIBRA-PKMzeta interaction. Starting from the known PKMzeta binding fragment, KIBRA residues 953-996, we will find the minimal binding site. Our data will guide design of peptides that bind PKMzeta that JK will inject into rat brain and test for memory and behaviour effects. Our studies will synergise with JK's parallel cell and in vivo studies towards detailed understanding and modulation of KIBRA function in memory.

Summary

KIBRA and the molecular mechanism of memory Our memories define who we are. Understanding the mechanisms behind the acquisition, storage and recall of memories is consequently a neuroscience holy grail. Memory depends on the interplay of different types of proteins (e.g. receptors, channels, enzymes, scaffold proteins). We will study KIBRA, a scaffold protein that is important for memory and that is linked to Alzheimer's disease. Since many details of how KIBRA functions in neurons (brain cells) are lacking, we will use experimental and computational methods to study KIBRA interactions with partner proteins and membranes (permeable biological boundaries between different parts of a cell or between cells). PKMzeta is crucial for long term memory storage The mechanisms of short and long term memory differ. The pivotal player in long term memory storage is a protein called PKMzeta which works by modifying other proteins. Modification by PKMzeta of receptor proteins, especially AMPA receptors, at the surface of neurons, for example, causes the receptors to move to the postsynaptic membrane where they contribute to electrical or chemical signalling between neurons to maintain memories. PKMzeta acts like a conveyor belt to carry AMPA receptors to the synapse. Modulation of PKMzeta activity can disrupt or enhance memory. KIBRA-PKMzeta interaction is vital for PKMzeta function Our collaborators have shown that KIBRA interaction with PKMzeta is crucial for PKMzeta's conveyor belt action. We will define details of KIBRA-PKMzeta interaction, e.g. the nature of the interface between the proteins, role of protein movement, and which protein components are most important for the interaction. Other KIBRA interactions that are important for memory We will study two other KIBRA interactions: with the key neuronal proteins Dendrin and Synaptopodin that are organisers of the molecular skeleton that gives neurons their characteristic shape; and KIBRA's interaction with membranes that maintains KIBRA in the correct location within neurons. Different parts of KIBRA are involved in its interactions with PKMzeta, Dendrin and Synaptopodin, and membranes. Methods We will use the complementary characteristics of multiple methods. NMR exploits the magnetic properties of nuclei to provide information about the shapes, shape changes and interactions of proteins at many locations throughout proteins and across a wide range of timescales (picoseconds to minutes). X-ray crystallography provides higher resolution shape information but less insight into dynamic behaviour. Other methods tell us about the strength and dynamics of protein interactions with other molecules. Computer simulations of protein behaviour provide insights that are not available from experiment alone, aid interpretation of experimental data and inform design of new experiments. Combining our study of molecules with our collaborator Joachim Kremerskothen's parallel experiments on cells and animals should lead to deep insight into KIBRA function. Long term goal: treatments for addictions, phobias, stress and anxiety disorders, and memory decline We will use our results to guide alterations in KIBRA that disrupt/enhance its interactions. Dr Kremerskothen will study how these KIBRA alterations affect neuron functions and memory processes in animals. This could help in the development of new molecules that affect memory. One existing molecule called ZIP that inhibits PKMzeta, for example, erases all long term memories yet permits formation of new memories. Improved understanding of memory-related molecular interactions such as those of KIBRA could help to develop molecules that specifically disrupt individual long term memories rather than all long term memories, for example to treat addiction, phobias, stress and anxiety disorders, or develop molecules that enhance memory performance in the elderly, Alzheimer's patients, or physical trauma victims.

Impact Summary

The research fits with BBSRC's Bioscience Underpinning Health priority and BBSRC Grand Challenge 3 - Fundamental bioscience enhancing lives and improving wellbeing (BBSRC Delivery Plan 2011-15). The research has benefit and interest in several areas, particularly due to its potential to contribute to ongoing research to find new treatments for memory-related afflictions such as addiction, phobias, stress and anxiety disorders, and memory decline. Economic and social burden of addictions (e.g. smoking, alcohol, illegal drugs, gambling) Addictions exert a huge socio-economic burden. In 2005, for example, 109000 UK deaths were attributable to smoking with £5.2bn NHS costs. ("The burden of smoking-related ill health in the UK", Tobacco Control 18 (2009) 262). Global alcohol-related costs were estimated at $210-665 billion in 2002. ("The global economic burden of alcohol", Drug Alcohol Rev. 25 (2006) 537). Research reported in The Independent (Feb 2002) showed that "drug addicts cost the NHS, the state benefits system and the criminal justice system around £6.8bn a year. The social costs of drug addiction, mainly the cost to victims of crime, amount to a further £12bn annually." Economic and social burden of stress and anxiety disorders, phobias and memory loss PTSD affects 2.6-10% of British soldiers deployed to Afghanistan and Iraq (669-2548 soldiers per year; Select Committee on Defence) and up to 30% of road accident victims. In the USA, anxiety disorders cost more than $42 billion annually ("The economic burden of anxiety disorders," J. Clin. Psych. 60, 1999) and the National Institute of Mental Health reported in 2005 that 8.7-18.1% of Americans suffer from phobias. Memory loss due to age, disease and trauma exerts a huge burden; e.g. the Dementia 2010 report (dementia2010.org) was headlined "Dementia costs UK plc £23 billion a year". This cost will rise as the proportion of UK society living beyond 65 increases rapidly. Pharmaceutical and biotech sector There is ongoing academic and industry research into memory modulator compounds e.g. propranolol (Nat. Neurosci. 12 (2009) 256). Zeta inhibitory peptide (ZIP; 13 residues) that inhibits PKMzeta can permanently erase all long term memories without preventing new memory formation (Science 317 (2007) 951). PKMzeta overexpression, moreover, enhances memory (Science 331 (2011) 1207). In 10-20 years, detailed understanding of memory-related molecular interactions such as those of KIBRA could be used by pharmaceutical/biotech/spin-out companies to develop biological or chemical molecules that specifically disrupt individual memories, for example to treat addiction, stress and anxiety disorders, or phobias, and molecules that enhance memory to treat memory decline due to ageing, disease or trauma. Public sector and general public The quality of life of many people would be improved by new drugs to treat memory-related afflictions such as addiction, stress and anxiety disorders, phobias and memory decline. Such treatments could, for example, significantly reduce NHS costs, reduce addiction-associated crime and other social problems, help military PTSD sufferers, and allow more people to enjoy a healthy old age. In the much shorter term, local schools and interested public will benefit from the proposed outreach activities (Pathways to Impact). People: lab and transferable skills The PDRA and PhD/undergrad students will benefit from substantial training from PIs and collaborators who have diverse expertise and interests, and from access to state-of-the-art equipment, for example national NMR facilities, local X-ray facilities, Diamond Synchrotron, and Oxford Protein Production Facility. The PDRA and students will also acquire transferable skills both from working on the project and through attending courses e.g. project management (including budgetary aspects), written and oral communication skills including public and media engagement, and enterprise.

Committee	Research Committee D (Molecules, cells and industrial biotechnology)
Research Topics	Neuroscience and Behaviour, Structural Biology
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search