Project 2: Tau uptake mechanisms and neuronal excitability in FTD

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Principal Investigator: KENNETH Stephen KOSIK
Organization: WEILL MEDICAL COLL OF CORNELL UNIV
Fiscal Year: 2020
Award: $296,067
Funding agency: National Institute of Neurological Disorders and Stroke

PROJECT SUMMARY/ABSTRACT — PROJECT 2 
 
Many unknown pathogenic steps lie between normal tau protein in a healthy neuron and the cellular 
dysfunction that occurs as a result of tau uptake and aggregation. The aims address two emerging facets of 
the pathobiology: the mechanism of tau uptake in neurons and astrocytes and the effects of tau inclusions on a 
set of functional cellular parameters. Tau uptake will be studied using a highly novel and highly collaborative 
approach called CRISPRi to identify in a non­biased manner genes involved in tau uptake followed by 
functional studies, which will validate the CRISPRi hits. A more focused approach to uptake will focus on 
HSPGs with a GAG microarray panel to identify HSPG chain features to which tau can bind specifically. Once 
tau is inside the cell we ask what are the effects of tau inclusions or tau mutations in human iPSC­derived 
neurons. We have synthesized multiple types of tau aggregates and studied their uptake conditions. A variety 
of techniques will be applied to pinpoint cellular defects due to the burden of tau inclusions. These techniques 
fall into two categories—expression sequencing and electrophysiological assessment. With regard to the 
former, the strength of our proposal is the use of single cell RNAseq to identify precisely gene expression 
changes in cells with tau inclusions compared to its neighbors that do not have tau inclusions. A further 
strength is the data that supports our hypothesis concerning the role of tRNA in triggering tau conformational 
changes that may lead to aggregation as well as an investigation of the role of cellular stress in inducing tRNA 
cleavage and the formation of half tRNAs known as tiRNAs. We have also hypothesized that tau can induce 
electrophysiologic dysfunction and over the past three years in collaboration with the physics department have 
built tools capable of detecting conduction deficits, alterations in the action potential at the axonal initial 
segment and hyer­excitability. The detection methods utilize a multi­electrode array (MEA) platform, modified 
MEAs that are patterned to confine neuron growth and direct signaling between neuronal ensembles, analytical 
tools for spike trains, and an automated cell harvesting device. The delineation of the effects on tau mutations 
and tau inclusions on cultured cells is a cellular phenotype that is not well­described in the field, but will be 
necessary in the future for small molecule screens. Working inter­changeably with several cellular systems will 
allow us to select the ideal context for each experimental question posed and explore multiple facets in a 
search for tau­related cellular phenotypes. This proposal rests on strong interactions among all the team 
members. 

Terms: <Action Potentials><Address><Affect><Anticodon><Astrocytes><Astrocytus><Astroglia><Axon><Back><Binding><CRISPR interference><CRISPRi><California><Categories><Cell Body><Cell Communication and Signaling><Cell Line><Cell Signaling><CellLine><Cells><Cellular Stress><Characteristics><Clustered Regularly Interspaced Short Palindromic Repeats interference><Collaborations><Collection><Computer software><Cultured Cells><Data><Defect><Dendrites><Detection><Devices><Dorsum><Dysfunction><Electrophysiology><Electrophysiology (science)><Enzyme Gene><Enzymes><Event><Functional disorder><Future><GAG><GAG Gene><Gametes><Gene Expression><Generalized Growth><Genes><Germ Cells><Germ-Line Cells><Glycans><Growth><HSPG><Harvest><Heparan Sulfate Proteoglycan><Human><Institutes><Intracellular Communication and Signaling><Investigation><Label><Lead><Link><MT-bound tau><Measures><Mediating><Methods><Microarray Analysis><Microarray-Based Analysis><Modern Man><Molecular Interaction><Msec><Nerve Cells><Nerve Unit><Neural Cell><Neurocyte><Neurons><Neurophysiology / Electrophysiology><Non-Polyadenylated RNA><Pathogenicity><Pathologic><Pathology><Pathway interactions><Pattern><Pb element><Phenotype><Physics><Physiopathology><Polysaccharides><Population><Preparation><Process><Proteoheparan Sulfate><RNA><RNA Gene Products><RNA Seq><RNA sequencing><RNAseq><Reproductive Cells><Resolution><Rest><Ribonucleic Acid><Role><Sex Cell><Signal Transduction><Signal Transduction Systems><Signaling><Small RNA><Software><Strains Cell Lines><Stress><System><Techniques><Technology><Testing><Time><Tissue Growth><Training><Transfer RNA><Translations><Triplet Codon-Amino Acid Adaptor><abnormal tau><abnormally aggregated tau protein><analytical tool><astrocytic glia><base><biological signal transduction><cell stress><collaborative approach><cultured cell line><design><designing><differential expression><differentially expressed><electrophysiological><falls><filamentous tau inclusion><heavy metal Pb><heavy metal lead><iPS><iPSC><iPSCs><induced pluripotent stem cell><initial cell><instrumentation><interest><member><microarray analyses><microarray technology><microtubule associated protein tau aggregation><microtubule associated protein tau deposit><microtubule associated protein tau mutation><microtubule bound tau><microtubule-associated protein tau mutation><microtubule-bound tau><millisecond><multi-electrode arrays><multielectrode arrays><mutant tau><mutation in microtubule associated protein tau><mutation in microtubule-associated protein tau><neural><neuronal><neuronal excitability><novel><ontogeny><paired helical filament of tau><pathogenic tau><pathogenic tau gene mutation><pathological change in tau><pathophysiology><pathway><programs><protein homeostasis><proteostasis><relating to nervous system><scRNA-seq><self-aggregate tau><sexual cell><single cell RNA-seq><single cell RNAseq><single-cell RNA sequencing><small molecule><social role><stress granule><tRNA><tau><tau PHF><tau Proteins><tau abnormality><tau accumulation><tau aggregate><tau aggregation><tau conformation><tau factor><tau fibrillization><tau filament><tau intronic mutation><tau mutation><tau neurofibrillary tangle><tau oligomer><tau paired helical filament><tau pathological change><tau polymerization><tau-tau interaction><tool><transcriptional differences><transcriptome sequencing><transcriptomics><transfer Ribonucleic acids><uptake><τ Proteins><τ aggregation><τ conformation><τ mutation>