Project description:Tau protein has critical roles in Alzheimer’s disease (AD) pathogenesis and neuronal excitation. Among tau gene mutations, A152T is reported to increase the risk of AD and neuronal excitability in mouse models. To examine the effects of tau gene mutations on neuronal excitability in human neurons, we introduced A152T or P301S mutation to induced pluripotent stem cells (iPSCs) using genome editing technology. To examine the effects of tau expression itself, we generated tau knockout or tau conjugate with a fluorescent protein. In excitatory neuronal culture, we found that the A152T mutation increases spontaneous neuronal excitation and the association of tau and Fyn. NMDAR antagonist blocked neuronal excitation in both the control and A152T neurons, indicating that the A152T mutation enhanced the intrinsic function of tau. Transcriptome analysis revealed structural changes by the A152T mutation. These data showed that the A152T tau gene mutation increases neuronal excitability through the tau-FYN-NMDAR pathway in excitatory neurons.
Project description:Tau is a microtubule-binding protein expressed in neurons and the equal ratio 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 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 tauopathypatients 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 to the adult brain in health and disease.
Project description:To explore the mechanistic basis of ApoE Ɛ4 vs ApoE Ɛ3 protein expression on endocytic pathways responsible for tau uptake in neurons and astrocytes and the maturation of neuronal networks, we have developed genotype matched co-cultures of iPSC derived astrocytes and neurons derived from isogenic triads of iPSC lines generated by the ADAPTED consortium (Schmid et al., 2019, 2020) . We show that isogenic iPSC derived APOE-E4 expressing astrocytes take up less extracellular tau than APOE-E3 or APOE null astrocytes, while isogenic neurons in monoculture take up equivalent amounts of tau primarily through macropinocytosis. Co-culture of neurons with genotype matched astrocytes increases the general uptake capacity of neurons by increasing the dependence of neuronal endocytosis on dynamin mediated pathways. Co-culture also enhances the emergence of spontaneous neuronal activity; however, the emergence of synchronous network activity is impaired by the expression of APOE-E4 genotype. We performed RNA-Seq on the astrocytes, neurons and co-cultures to understand the molecular pathway changes associated with different ApoE genotypes.
Project description:Tauopathies are age-associated neurodegenerative diseases whose mechanistic underpinnings remain elusive, partially due to lack of appropriate human models. Here, we engineered new human induced pluripotent stem cell (hiPSC)-derived neuronal lines to express 4R Tau and 4R Tau carrying the P301S MAPT mutation when differentiated into neurons. 4R-P301S neurons display progressive Tau inclusions upon seeding with Tau fibrils and recapitulate features of tauopathy phenotypes including shared transcriptomic signatures, autophagic body accumulation, and reduced neuronal activity. A CRISPRi screen of genes associated with Tau pathobiology identified over 500 genetic modifiers of seeding-induced Tau propagation, including retromer VPS29 and genes in the UFMylation cascade. In progressive supranuclear palsy (PSP) and Alzheimer’s Disease (AD) brains, the UFMylation cascade is altered in neurofibrillary-tangle-bearing neurons. Inhibiting the UFMylation cascade in vitro and in vivo suppressed seeding-induced Tau propagation. This model provides a robust platform to identify novel therapeutic strategies for 4R tauopathy.
Project description:The Tau (MAPT) protein drives neuronal dysfunction and toxicity in the brain in Alzheimer’s disease (AD) and other Tauopathies. To dissect the complexity of the Tau interactome that underlies this process we used two proteomic approaches to characterize the dynamic and multifaceted nature of Tau protein-protein interactions in human induced pluripotent stem cell (iPSC)-derived neurons. We used engineered ascorbic acid peroxidase (APEX) for spatiotemporally restricted mapping of Tau interaction proteins in combination with quantitative affinity purification mass spectrometry (AP-MS) to interrogate disease-related changes in the Tau interactome. The APEX method resolved subcellular interactions of wild-type Tau at amino acid level resolution in living neurons as well as novel activity-dependent interactions of Tau with presynaptic vesicle proteins that occurred during Tau secretion from neurons. Among the many Tau interacting proteins revealed by AP-MS, the interaction of mitochondrial proteins with wild-type Tau (TauWT) was more robust than with TauV337M or TauP301L. The mitochondrial proteins that preferentially interacted with TauWT comprised a protein module that is downregulated in multi-omics analyses of human AD brains. Mitochondrial bioenergetics were altered in TauV337M compared to TauWT human iPSC-derived neurons confirming the impact of Tau on mitochondria. These Tau interactome analyses open up novel disease-related processes as potential therapeutic targets to block Tau-mediated pathogenesis.
Project description:Purpose: The goal of this study was to assess gene expression changes upon SAHA treatment in neurons derived from patients with A152T Tau mutation
Project description:Aggregation of the microtubule-associated protein, tau, can lead to neurofibrillary tangle formation in neurons and glia, the hallmark of tauopathy. The cellular damages induced by the tau overexpression and aggregation may lead to multiple pathologic features of tauopathy. However, the effect of aging on tauopathy has not been elucidated yet. Here, we conducted lncRNA/mRNA sequencing analysis on P301S mutant Tau transgenic mouse model (PS19) with different ages to track the genetic changes occurred by the aging and progression of tau overexpression.
Project description:Tauopathies are age-associated neurodegenerative diseases whose mechanistic underpinnings remain elusive, partially due to lack of appropriate human models. Here, we engineered new human induced pluripotent stem cell (hiPSC)-derived neuronal lines to express 4R Tau and 4R Tau carrying the P301S MAPT mutation when differentiated into neurons. 4R-P301S neurons display progressive Tau inclusions upon seeding with Tau fibrils and recapitulate features of tauopathy phenotypes including shared transcriptomic signatures, autophagic body accumulation, and reduced neuronal activity. A CRISPRi screen of genes associated with Tau pathobiology identified over 500 genetic modifiers of seeding-induced Tau propagation, including retromer VPS29 and genes in the UFMylation cascade. In progressive supranuclear palsy (PSP) and Alzheimer’s Disease (AD) brains, the UFMylation cascade is altered in neurofibrillary-tangle-bearing neurons. Inhibiting the UFMylation cascade in vitro and in vivo suppressed seeding-induced Tau propagation. This model provides a robust platform to identify novel therapeutic strategies for 4R tauopathy.