2021 |
Mitchison, Timothy J [⬀] Song, Yuyu |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
Nuclear Transport as a Molecular and Cellular Vulnerability in Ad
Abstract Molecular trafficking between the nucleus and the cytoplasm is essential for cellular health and is tightly regulated in all cell types including those in the brain. Recent publications demonstrated nuclear transport defects in neurons in Alzheimer?s disease (AD) and related dementias (ADRDs). AD/ADRDs are caused, in part, by misfolded tau protein, suggesting misfolded tau may impair nuclear transport by aberrantly interacting with nuclear pore proteins. We propose that late-onset neurodegenerative disease, such as AD, reflects two vulnerabilities: (i) At the cellular level, intrinsic loss of nuclear import efficiency during the neural differentiation program sensitizes the neurons to damages associated with misfolded tau. (ii) At the molecular level, nuclear pore complexes (NPCs) are selectively vulnerable to disruption by misfolded proteins because their activity depends on exposed hydrophobic phenylalanine-glycine (FG) repeats that are easily disrupted by misfolded AD/ADRD-tau and turn over very slowly. To test these hypothesis, we developed novel optogenetic nuclear transport assays, based on photo-activatable NLS/NES elements. We now propose to combine Mitchison group?s expertise in advanced microscopy and image analysis with Song group?s expertise in neuron cell biology and pathology models to measure rates of nuclear import and export in living neurons and test the effects of neural differentiation, misfolded tau, and drug candidates that may alleviate the effects of misfolded tau on nuclear transport. We will (i) characterize the change in nuclear transport rates during neural differentiation, (ii) compare the sensitivity of nuclear transport to AD/ADRD-related misfolded tau challenges (e.g. G272V-tau and P301S-tau) in neurons and neural progenitors, and (iii) investigate the underlying molecular mechanisms using super-resolution microscopy and immunoassays. The transport assays will also enable future translational programs aimed at rescuing nuclear transport in aging neurons. As a test case, we will characterize drugs that increase O-linked b-N-acetylglucosamine (O-GlcNAc) modification of intracellular proteins. This modification is thought to inhibit aggregation of misfolded proteins such as tau. However, FG repeat in NPC are among the most O-GlcNAc modified proteins. We propose that the function of this druggable modification is to protect the intrinsic vulnerability of NPCs to damage by misfolded proteins. Success on this R21 pilot will set the stage for moving our optical reporter strategy into mouse models of brain aging and degeneration, and for identifying drug targets and testing candidate therapeutic molecules in high- content assay formats.
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