Functional Role of Presynaptic Tau During Aging
Lead Research Organisation:
King's College London
Department Name: Neuroscience
Abstract
The functional significance of tau protein in the presynapse in unknown. Recent studies indicate that tau interacts with the synaptic vesicle protein synaptogyrin3, and that this interaction affects synaptic vesicle mobility, which in turn modulates neurotransmission. Tau is also known to be released from neurons in an activity-dependent manner. Importantly, abnormal tau protein in the presynapse affects both tau's interaction with synaptic vesicles and its ability to be released upon stimulation. The aim of this project is to further investigate the function of tau in the presynapse by using the organotypic slice model, which is advantageous for several reasons: 1. Synaptic and anatomical connections are retained, 2. all brain cells are present, 3. Cultures can be maintained long-term (up to 6 months) allowing synapses to mature, and 4. Slice cultures show accelerated aging making it feasible to study age-related changes to synapses.
The first experimental aim is to compare presynaptic tau function in slices from both wild-type and tau transgenic mice (htau), which exhibit accelerated aging phenotypes, at multiple time points. The amount of tau present on synaptic vesicles and filamentous actin levels will be quantified using imaging-based assays. Synaptic vesicle mobility will also be assessed with established fluorescent probes (i.e., FM dyes). The second project aim is to determine the effects on synapse health of blocking the tau-synaptogyrin-3 interaction. This can be achieved in slices by AAV-mediated viral delivery of shRNA to knockdown synaptogyrin-3. Thirdly, the amount of tau released by cultures will be measured using an ELISA-based approach, and comparisons will be made between wild-type and transgenic cultures, with and without synaptogyrin-3 knockdown. Finally, biochemical assays will be used to elucidate the exact binding residues within tau and synaptogyrin-3 that mediate the interaction. Mutagenesis will be performed within the N-terminus of tau and the cytoplasmic regions of synaptogyrin-3, and purified fusion proteins will be used in pulldown assays.
Tau is a widely-studied protein due to its causal role in neurodegenerative disease, yet the functional significance of normal and abnormal tau in the synapse is unclear. While some research into post-synaptic tau function exists, less is known about the role of tau in the presynapse, and how and why tau is released upon neuronal stimulation.
The first experimental aim is to compare presynaptic tau function in slices from both wild-type and tau transgenic mice (htau), which exhibit accelerated aging phenotypes, at multiple time points. The amount of tau present on synaptic vesicles and filamentous actin levels will be quantified using imaging-based assays. Synaptic vesicle mobility will also be assessed with established fluorescent probes (i.e., FM dyes). The second project aim is to determine the effects on synapse health of blocking the tau-synaptogyrin-3 interaction. This can be achieved in slices by AAV-mediated viral delivery of shRNA to knockdown synaptogyrin-3. Thirdly, the amount of tau released by cultures will be measured using an ELISA-based approach, and comparisons will be made between wild-type and transgenic cultures, with and without synaptogyrin-3 knockdown. Finally, biochemical assays will be used to elucidate the exact binding residues within tau and synaptogyrin-3 that mediate the interaction. Mutagenesis will be performed within the N-terminus of tau and the cytoplasmic regions of synaptogyrin-3, and purified fusion proteins will be used in pulldown assays.
Tau is a widely-studied protein due to its causal role in neurodegenerative disease, yet the functional significance of normal and abnormal tau in the synapse is unclear. While some research into post-synaptic tau function exists, less is known about the role of tau in the presynapse, and how and why tau is released upon neuronal stimulation.
People |
ORCID iD |
| Wendy Noble (Primary Supervisor) | |
| Emily Groves (Student) |