Evidences p(HGNC:MAPT, var("p.Pro301Ser")) to p(HGNC:MAPT, var("p.Pro301Ser"))

After extensive washing, monomeric and aggregated tau-Dylight were both detected within cells expressing the neuron-specific microtubule-associated protein MAP2, confirming that both forms of tau enter neurons

Tau-Dylight was found predominantly within the somatic compartment of neurons (Figure 1A)

After a 4-hr incubation with extracellular tau, flow cytometry analysis (Figures 1B and 1C) revealed that 83% and 73% of dissociated cells contained monomeric or aggregated tau-Dylight, respectively, demonstrating that extracellular tau efficiently enters human neurons in culture

Internalization of monomeric tau (P301S) and wild-type tau was comparable and concentration dependent (Figure S3A), confirming that the P301S mutation does not confer the ability to efficiently enter neurons, nor is this form of tau likely to aggregate in extracellular media during the 3- to 4-hr incubation period

Incubation of tau-pHrodo with human neurons at a range of concentrations from 2.5 to 25 nM (0.12–1.2 µg.mL-1, diluted in culture medium) showed that tau entry to neurons is rapid, as visualized by live imaging

We found that FLAG-tagged tau enters neurons efficiently and that internalized tau persists at detectable levels within neurons for at least 4 days (Figure S4A)

Intracellular fluorescent punctae were observed within the first 10 min of exposure of neurons to monomeric tau-pHrodo (Figure 2A; Video S1). Tau-pHrodo-positive structures increased in size and intensity over the 4-hr course of the assay. These structures were present within neurites and accumulated in the cell bodies of neurons

In the presence of 15 and 25 nM monomeric tau-pHrodo, the number of tau-pHrodo-positive objects approached a plateau (>90% of final measurement) after approximately 1 hr (Figure 2C)

These kinetics of aggregated tau-pHrodo entry are similar to that of both lower concentrations of monomeric tau (2.5 nM) and of low-molecular weight (10-kDa) dextran-pHrodo (same molarity as monomeric tau samples; Figures S5A–S5C

In agreement with the monomeric tau-pHrodo experiments, monomeric tau-Dylight was rapidly taken up into neurons (Figure 3C

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If you find BEL Commons useful in your work, please consider citing: Hoyt, C. T., Domingo-Fernández, D., & Hofmann-Apitius, M. (2018). BEL Commons: an environment for exploration and analysis of networks encoded in Biological Expression Language. Database, 2018(3), 1–11.