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  • Recapitulation of endogenous 4R tau expression and formation of insoluble tau in directly reprogrammed human neurons.

Recapitulation of endogenous 4R tau expression and formation of insoluble tau in directly reprogrammed human neurons.

Cell stem cell (2022-06-07)
Lucia S Capano, Chihiro Sato, Elena Ficulle, Anan Yu, Kanta Horie, Ji-Sun Kwon, Kyle F Burbach, Nicolas R Barthélemy, Susan G Fox, Celeste M Karch, Randall J Bateman, Henry Houlden, Richard I Morimoto, David M Holtzman, Karen E Duff, Andrew S Yoo
ABSTRACT

Tau is a microtubule-binding protein expressed in neurons, and the equal ratios between 4-repeat (4R) and 3-repeat (3R) isoforms are maintained in normal adult brain function. Dysregulation of 3R:4R ratio causes tauopathy, and human neurons that recapitulate tau isoforms in health and disease will provide a platform for elucidating pathogenic processes involving tau pathology. We carried out extensive characterizations of tau isoforms expressed in human neurons derived by microRNA-induced neuronal reprogramming of adult fibroblasts. Transcript and protein analyses showed that miR neurons expressed all six isoforms with the 3R:4R isoform ratio equivalent to that detected in human adult brains. Also, miR neurons derived from familial tauopathy patients with a 3R:4R ratio altering mutation showed increased 4R tau and the formation of insoluble tau with seeding activity. Our results collectively demonstrate the utility of miRNA-induced neuronal reprogramming to recapitulate endogenous tau regulation comparable with the adult brain in health and disease.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
DAPT, ≥98% (HPLC), solid
Sigma-Aldrich
Anti-Microtubule-Associated Protein 2 (MAP2) Antibody, Chemicon®, from rabbit
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cOmplete, Mini Protease Inhibitor Cocktail, Tablets provided in a glass vial
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N6,2′-O-Dibutyryladenosine 3′,5′-cyclic monophosphate sodium salt, ≥96% (HPLC), powder
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Fibronectin bovine plasma, powder, BioReagent, suitable for cell culture
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Laminin from Engelbreth-Holm-Swarm murine sarcoma basement membrane, 1-2 mg/mL in Tris-buffered saline, 0.2 μm filtered, BioReagent, suitable for cell culture
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Valproic Acid, Sodium Salt, A cell-permeable, short-chained fatty acid that inhibits histone deacetylase (IC₅₀ = 400 µM for HDAC1).