LRP1 is a master regulator of tau uptake and spread

Abstract

The spread of protein aggregates during disease progression is a common theme underlying many neurodegenerative diseases. The microtubule-associated protein tau has a central role in the pathogenesis of several forms of dementia known as tauopathies—including Alzheimer’s disease, frontotemporal dementia and chronic traumatic encephalopathy1. Progression of these diseases is characterized by the sequential spread and deposition of protein aggregates in a predictable pattern that correlates with clinical severity2. This observation and complementary experimental studies3,4 have suggested that tau can spread in a prion-like manner, by passing to naive cells in which it templates misfolding and aggregation. However, although the propagation of tau has been extensively studied, the underlying cellular mechanisms remain poorly understood. Here we show that the low-density lipoprotein receptor-related protein 1 (LRP1) controls the endocytosis of tau and its subsequent spread. Knockdown of LRP1 significantly reduced tau uptake in H4 neuroglioma cells and in induced pluripotent stem cell-derived neurons. The interaction between tau and LRP1 is mediated by lysine residues in the microtubule-binding repeat region of tau. Furthermore, downregulation of LRP1 in an in vivo mouse model of tau spread was found to effectively reduce the propagation of tau between neurons. Our results identify LRP1 as a key regulator of tau spread in the brain, and therefore a potential target for the treatment of diseases that involve tau spread and aggregation.

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Fig. 1: Tau uptake is mediated by LRP1.
Fig. 2: Tau interacts with LRP1 in a similar manner to known ligands.
Fig. 3: LRP1 mediates tau uptake in neurons.
Fig. 4: LRP1 knockdown reduces tau spread in vivo.

Data availability

Source data are available for graphs plotted in Figs. 14 and Extended Data Figs. 13. Scans of the full western blot gels can be found in Supplementary Fig. 1. All other data are available from the corresponding author upon reasonable request.

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Acknowledgements

This study was funded by the National Institutes of Health (NIH), K99 AG064116 (J.N.R.), DP2 GM119139 (M.K.), R01 AG062359 (M.K.), R56 AG057528 (M.K.), U54 NS 100717 (M.K., K.S.K.), Tau Consortium (M.K., K.S.K.), German Center for Neurodegenerative Diseases (S.W.), Ben Barres Early Career Acceleration Award from the Chan Zuckerberg Initiative (M.K.), Tri-counties Blood Bank (J.N.R.), Dr Miriam and Sheldon G. Adelson Medical Research Foundation (K.S.K.), Larry L. Hillblom Foundation (K.S.K.) and Edward N. & Della L. Thome Memorial Foundation (K.S.K.). We thank the Laboratory for Stem Cell Biology and Engineering at UCSB for use of their flow cytometer, the Neuroscience Research Institute Microscopy Facility for use of their microscopes, J. Dong for help in the early stages of this project and P. Davies for providing the MC-1 antibody.

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Contributions

J.N.R. and K.S.K. conceived the study. J.N.R. and K.S.K. designed the study. J.N.R. performed most of the experiments and analysed the data, assisted by G.L., E.G., M.A., C.C., Y.E.S., C.L. and I.H. C.C., V.G., S.W. and B.T.H. provided AAVs used in the study. M.K. provided CRISPRi cell lines. J.N.R. and K.S.K. wrote the manuscript, and all authors discussed the results and commented on the manuscript.

Corresponding author

Correspondence to Kenneth S. Kosik.

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K.S.K. is on the Board of Directors of the Rainwater Charitable Trust.

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Peer review information Nature thanks Katrin Deinhardt, Joachim Herz and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data figures and tables

Extended Data Fig. 1 Tau uptake is regulated by LRP1.

a, Gating strategy for tau uptake assay. First, cells were gated on forward scatter/side scatter (FSC/SSC; mean FSC-A, around 8,000,000; mean SSC-A, around 800,000). Cells were then gated on forward scatter height (FSC-H) versus width to discriminate doublets. Dead cells were removed from the analysis using propidium iodide as a stain, and positive cells were determined by gating on a negative (no tau added) population. b, Internalization controls for tau uptake assay (n = 6). c, Quantitative PCR analysis of various genes with sgRNA in H4i cells. The first column represents the NT sgRNA control for each target (n = 3). d, Uptake of phosphorylated (p2N4R) or mutated (2N4RP301L) full-length tau in H4i cells (n = 8). e, Western blot analysis of LRP1 in wild-type, NT sgRNA or LRP1 sgRNA H4i cells. All results in be were obtained in three independent experiments and normalized to wild-type uptake (100%). Data are expressed as mean ± s.d. with individual data points shown. One-way ANOVA with Tukey’s method, two-sided was performed to determine significance; ****P < 0.0001. Source Data

Extended Data Fig. 2 LRP1 ligands influence tau uptake.

a, Uptake of 2N4R tau with competition from RAP or lysine-capped RAP. b, Uptake of 2N4R tau with competition from RAP or mtRAP. c, Corrected integrated density of surface HA staining for different ectodomain cell lines (n = 60). d, Co-immunoprecipitation of HA–mLRP4 with myc–2N4R (n = 3). e, Uptake of various ligands at indicated concentrations (1 h). f, Western blot analysis of conditioned medium (CM) from HEK293T cells, mock-treated or overexpressing ApoE (n = 3). g, Uptake of full-length tau and transferrin in conditioned medium from f (n = 9). All experiments in a, b, e were performed in biological duplicate over three independent experiments (n = 6) with representative experiments shown. Data are expressed as mean ± s.d. with individual data points shown. One-way ANOVA with Tukey’s method, two-sided was performed to determine significance; ****P < 0.0001. Source Data

Extended Data Fig. 3 LRP1 expression influences tau spread.

a, Quantitative PCR analysis of LRP1 expression in iPSNs (n = 3; P < 0.0001). b, Quantitative PCR analysis of LRP1 expression from mouse cortex transduced with human synapsin (hSyn) scramble shRNA or LRP1 shRNA (n = 3; P = 0.0412). c, Quantification of tau spread in mice broken down by sex (males n = 4, females n = 3; two-way ANOVA, sex = NS, P = 0.5335). d, Immunofluorescence of mouse primary culture transduced with AAVmRuby-hSyn-shLRP1 (green, Tau; red, mRuby; blue, Hoechst; scale bar, 20 μm, n = 3). d, Immunohistochemistry of hTau+ astrocytes in mice injected with PBS (green, Sox2; red, hTau; blue, Hoechst; scale bar, 20 μm, n = 3). e, Immunohistochemistry of mice injected with scramble shRNA and LRP1 shRNA (green, LRP1; blue; Hoechst, scale bars, 20 μm; n = 3). Unpaired t-test, two-tailed was performed to determine statistical significance for quantitative PCR in a, b; *P ≤ 0.05, ****P < 0.0001. Source Data

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Rauch, J.N., Luna, G., Guzman, E. et al. LRP1 is a master regulator of tau uptake and spread. Nature 580, 381–385 (2020). https://doi.org/10.1038/s41586-020-2156-5

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