The cytoplasmic mislocalization and aggregation of TAR DNA-binding protein-43 (TDP-43) is a common histopathological hallmark of the amyotrophic lateral sclerosis and frontotemporal dementia disease spectrum (ALS/FTD). However, the composition of aggregates and their contribution to the disease process remain unknown. Here we used proximity-dependent biotin identification (BioID) to interrogate the interactome of detergent-insoluble TDP-43 aggregates and found them enriched for components of the nuclear pore complex and nucleocytoplasmic transport machinery. Aggregated and disease-linked mutant TDP-43 triggered the sequestration and/or mislocalization of nucleoporins and transport factors, and interfered with nuclear protein import and RNA export in mouse primary cortical neurons, human fibroblasts and induced pluripotent stem cell–derived neurons. Nuclear pore pathology is present in brain tissue in cases of sporadic ALS and those involving genetic mutations in TARDBP and C9orf72. Our data strongly implicate TDP-43-mediated nucleocytoplasmic transport defects as a common disease mechanism in ALS/FTD.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.


All prices are NET prices.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.


  1. 1.

    Ling, S. C., Polymenidou, M. & Cleveland, D. W. Converging mechanisms in ALS and FTD: disrupted RNA and protein homeostasis. Neuron 79, 416–438 (2013).

  2. 2.

    Neumann, M. et al. Ubiquitinated TDP-43 in frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Science 314, 130–133 (2006).

  3. 3.

    Igaz, L. M. et al. Enrichment of C-terminal fragments in TAR DNA-binding protein-43 cytoplasmic inclusions in brain but not in spinal cord of frontotemporal lobar degeneration and amyotrophic lateral sclerosis. Am. J. Pathol. 173, 182–194 (2008).

  4. 4.

    Buratti, E. et al. TDP-43 binds heterogeneous nuclear ribonucleoprotein A/B through its C-terminal tail: an important region for the inhibition of cystic fibrosis transmembrane conductance regulator exon 9 splicing. J. Biol. Chem. 280, 37572–37584 (2005).

  5. 5.

    Gitcho, M. A. et al. TDP-43 A315T mutation in familial motor neuron disease. Ann. Neurol. 63, 535–538 (2008).

  6. 6.

    Nonaka, T., Kametani, F., Arai, T., Akiyama, H. & Hasegawa, M. Truncation and pathogenic mutations facilitate the formation of intracellular aggregates of TDP-43. Hum. Mol. Genet. 18, 3353–3364 (2009).

  7. 7.

    Guo, W. et al. An ALS-associated mutation affecting TDP-43 enhances protein aggregation, fibril formation and neurotoxicity. Nat. Struct. Mol. Biol. 18, 822–830 (2011).

  8. 8.

    Li, Y. R., King, O. D., Shorter, J. & Gitler, A. D. Stress granules as crucibles of ALS pathogenesis. J. Cell Biol. 201, 361–372 (2013).

  9. 9.

    Fallini, C., Bassell, G. J. & Rossoll, W. The ALS disease protein TDP-43 is actively transported in motor neuron axons and regulates axon outgrowth. Hum. Mol. Genet. 21, 3703–3718 (2012).

  10. 10.

    Chou, C. C. et al. PABPN1 suppresses TDP-43 toxicity in ALS disease models. Hum. Mol. Genet. 24, 5154–5173 (2015).

  11. 11.

    Igaz, L. M. et al. Expression of TDP-43 C-terminal fragments in vitro recapitulates pathological features of TDP-43 proteinopathies. J. Biol. Chem. 284, 8516–8524 (2009).

  12. 12.

    Roux, K. J., Kim, D. I., Raida, M. & Burke, B. A promiscuous biotin ligase fusion protein identifies proximal and interacting proteins in mammalian cells. J. Cell Biol. 196, 801–810 (2012).

  13. 13.

    Kim, D. I. et al. Probing nuclear pore complex architecture with proximity-dependent biotinylation. Proc. Natl. Acad. Sci. USA 111, E2453–E2461 (2014).

  14. 14.

    Rout, M. P. et al. The yeast nuclear pore complex: composition, architecture, and transport mechanism. J. Cell Biol. 148, 635–651 (2000).

  15. 15.

    Lim, L., Wei, Y., Lu, Y. & Song, J. ALS-causing mutations significantly perturb the self-assembly and interaction with nucleic acid of the intrinsically disordered prion-like domain of TDP-43. PLoS Biol. 14, e1002338 (2016).

  16. 16.

    Wright, P. E. & Dyson, H. J. Intrinsically disordered proteins in cellular signalling and regulation. Nat. Rev. Mol. Cell Biol. 16, 18–29 (2015).

  17. 17.

    Halfmann, R., Wright, J. R., Alberti, S., Lindquist, S. & Rexach, M. Prion formation by a yeast GLFG nucleoporin. Prion 6, 391–399 (2012).

  18. 18.

    Frey, S. & Görlich, D. A saturated FG-repeat hydrogel can reproduce the permeability properties of nuclear pore complexes. Cell 130, 512–523 (2007).

  19. 19.

    Kato, M. et al. Cell-free formation of RNA granules: low complexity sequence domains form dynamic fibers within hydrogels. Cell 149, 753–767 (2012).

  20. 20.

    Lam, S. S. et al. Directed evolution of APEX2 for electron microscopy and proximity labeling. Nat. Methods 12, 51–54 (2015).

  21. 21.

    Ayala, Y. M., Misteli, T. & Baralle, F. E. TDP-43 regulates retinoblastoma protein phosphorylation through the repression of cyclin-dependent kinase 6 expression. Proc. Natl. Acad. Sci. USA 105, 3785–3789 (2008).

  22. 22.

    Broers, J. L., Ramaekers, F. C., Bonne, G., Yaou, R. B. & Hutchison, C. J. Nuclear lamins: laminopathies and their role in premature ageing. Physiol. Rev. 86, 967–1008 (2006).

  23. 23.

    Frost, B., Bardai, F. H. & Feany, M. B. Lamin dysfunction mediates neurodegeneration in tauopathies. Curr. Biol. 26, 129–136 (2016).

  24. 24.

    Crisp, M. et al. Coupling of the nucleus and cytoplasm: role of the LINC complex. J. Cell Biol. 172, 41–53 (2006).

  25. 25.

    Zhang, Q. et al. Nesprin-1 and -2 are involved in the pathogenesis of Emery Dreifuss muscular dystrophy and are critical for nuclear envelope integrity. Hum. Mol. Genet. 16, 2816–2833 (2007).

  26. 26.

    Yamashita, T., Aizawa, H., Teramoto, S., Akamatsu, M. & Kwak, S. Calpain-dependent disruption of nucleo-cytoplasmic transport in ALS motor neurons. Sci. Rep. 7, 39994 (2017).

  27. 27.

    Estes, P. S. et al. Motor neurons and glia exhibit specific individualized responses to TDP-43 expression in a Drosophila model of amyotrophic lateral sclerosis. Dis. Model. Mech. 6, 721–733 (2013).

  28. 28.

    Kinoshita, Y. et al. Nuclear contour irregularity and abnormal transporter protein distribution in anterior horn cells in amyotrophic lateral sclerosis. J. Neuropathol. Exp. Neurol. 68, 1184–1192 (2009).

  29. 29.

    Zhan, L., Hanson, K. A., Kim, S. H., Tare, A. & Tibbetts, R. S. Identification of genetic modifiers of TDP-43 neurotoxicity in Drosophila. PLoS One 8, e57214 (2013).

  30. 30.

    Cairns, N. J. et al. TDP-43 proteinopathy in familial motor neurone disease with TARDBP A315T mutation: a case report. Neuropathol. Appl. Neurobiol. 36, 673–679 (2010).

  31. 31.

    Perwitasari, O. et al. Antiviral efficacy of verdinexor in vivo in two animal models of influenza a virus infection. PLoS One 11, e0167221 (2016).

  32. 32.

    Tajiri, N. et al. A Nuclear attack on traumatic brain injury: sequestration of cell death in the nucleus. CNS Neurosci. Ther. 22, 306–315 (2016).

  33. 33.

    Zhang, K. et al. The C9orf72 repeat expansion disrupts nucleocytoplasmic transport. Nature 525, 56–61 (2015).

  34. 34.

    Grima, J. C. et al. Mutant huntingtin disrupts the nuclear pore complex. Neuron 94, 93–107.e6 (2017).

  35. 35.

    Soo, K. Y. et al. Rab1-dependent ER-Golgi transport dysfunction is a common pathogenic mechanism in SOD1, TDP-43 and FUS-associated ALS. Acta Neuropathol. 130, 679–697 (2015).

  36. 36.

    Alami, N. H. et al. Axonal transport of TDP-43 mRNA granules is impaired by ALS-causing mutations. Neuron 81, 536–543 (2014).

  37. 37.

    Nofrini, V., Di Giacomo, D. & Mecucci, C. Nucleoporin genes in human diseases. Eur. J. Hum. Genet. 24, 1388–1395 (2016).

  38. 38.

    Kaneb, H. M. et al. Deleterious mutations in the essential mRNA metabolism factor, hGle1, in amyotrophic lateral sclerosis. Hum. Mol. Genet. 24, 1363–1373 (2015).

  39. 39.

    Freibaum, B. D. et al. GGGGCC repeat expansion in C9orf72 compromises nucleocytoplasmic transport. Nature 525, 129–133 (2015).

  40. 40.

    Jovičić, A. et al. Modifiers of C9orf72 dipeptide repeat toxicity connect nucleocytoplasmic transport defects to FTD/ALS. Nat. Neurosci. 18, 1226–1229 (2015).

  41. 41.

    Zhang, Y. J. et al. C9ORF72 poly(GA) aggregates sequester and impair HR23 and nucleocytoplasmic transport proteins. Nat. Neurosci. 19, 668–677 (2016).

  42. 42.

    Pujol, G., Söderqvist, H. & Radu, A. Age-associated reduction of nuclear protein import in human fibroblasts. Biochem. Biophys. Res. Commun. 294, 354–358 (2002).

  43. 43.

    D’Angelo, M. A., Raices, M., Panowski, S. H. & Hetzer, M. W. Age-dependent deterioration of nuclear pore complexes causes a loss of nuclear integrity in postmitotic cells. Cell 136, 284–295 (2009).

  44. 44.

    Ward, M. E. et al. Early retinal neurodegeneration and impaired Ran-mediated nuclear import of TDP-43 in progranulin-deficient FTLD. J. Exp. Med. 211, 1937–1945 (2014).

  45. 45.

    Nishimura, A. L. et al. Nuclear import impairment causes cytoplasmic trans-activation response DNA-binding protein accumulation and is associated with frontotemporal lobar degeneration. Brain 133, 1763–1771 (2010).

  46. 46.

    Amlie-Wolf, A. et al. Transcriptomic changes due to cytoplasmic TDP-43 expression reveal dysregulation of histone transcripts and nuclear chromatin. PLoS One 10, e0141836 (2015).

  47. 47.

    Woerner, A. C. et al. Cytoplasmic protein aggregates interfere with nucleocytoplasmic transport of protein and RNA. Science 351, 173–176 (2016).

  48. 48.

    Gasset-Rosa, F. et al. Polyglutamine-expanded huntingtin exacerbates age-related disruption of nuclear integrity and nucleocytoplasmic transport. Neuron 94, 48–57.e4 (2017).

  49. 49.

    Lee, K. H. et al. C9orf72 dipeptide repeats impair the assembly, dynamics, and function of membrane-less organelles. Cell 167, 774–788.e17 (2016).

  50. 50.

    Molliex, A. et al. Phase separation by low complexity domains promotes stress granule assembly and drives pathological fibrillization. Cell 163, 123–133 (2015).

  51. 51.

    Ayala, Y. M. et al. Structural determinants of the cellular localization and shuttling of TDP-43. J. Cell Sci. 121, 3778–3785 (2008).

  52. 52.

    Estes, P. S. et al. Wild-type and A315T mutant TDP-43 exert differential neurotoxicity in a Drosophila model of ALS. Hum. Mol. Genet. 20, 2308–2321 (2011).

  53. 53.

    Joardar, A. et al. PPAR gamma activation is neuroprotective in a Drosophila model of ALS based on TDP-43. Hum. Mol. Genet. 24, 1741–1754 (2015).

  54. 54.

    Williams, K. R. et al. hnRNP-Q1 represses nascent axon growth in cortical neurons by inhibiting Gap-43 mRNA translation. Mol. Biol. Cell 27, 518–534 (2016).

  55. 55.

    Fallini, C., Bassell, G. J. & Rossoll, W. High-efficiency transfection of cultured primary motor neurons to study protein localization, trafficking, and function. Mol. Neurodegener. 5, 17 (2010).

  56. 56.

    Donnelly, C. J. et al. RNA toxicity from the ALS/FTD C9ORF72 expansion is mitigated by antisense intervention. Neuron 80, 415–428 (2013).

  57. 57.

    Rha, J. et al. The RNA-binding protein, ZC3H14, is required for proper poly(A) tail length control, expression of synaptic proteins, and brain function in mice. Hum. Mol. Genet. 26, 3663–3681 (2017).

  58. 58.

    Käll, L., Canterbury, J. D., Weston, J., Noble, W. S. & MacCoss, M. J. Semi-supervised learning for peptide identification from shotgun proteomics datasets. Nat. Methods 4, 923–925 (2007).

  59. 59.

    Kuo, L. J. & Yang, L. X. Gamma-H2AX – a novel biomarker for DNA double-strand breaks. In Vivo 22, 305–309 (2008).

  60. 60.

    Hatch, E. M., Fischer, A. H., Deerinck, T. J. & Hetzer, M. W. Catastrophic nuclear envelope collapse in cancer cell micronuclei. Cell 154, 47–60 (2013).

  61. 61.

    Caballero-Benítez, A. & Morán, J. Caspase activation pathways induced by staurosporine and low potassium: role of caspase-2. J. Neurosci. Res. 71, 383–396 (2003).

  62. 62.

    Soderholm, J. F. et al. Importazole, a small molecule inhibitor of the transport receptor importin-β. ACS Chem. Biol. 6, 700–708 (2011).

  63. 63.

    Fallini, C. et al. The survival of motor neuron (SMN) protein interacts with the mRNA-binding protein HuD and regulates localization of poly(A) mRNA in primary motor neuron axons. J. Neurosci. 31, 3914–3925 (2011).

  64. 64.

    Coyne, A. N. et al. Futsch/MAP1B mRNA is a translational target of TDP-43 and is neuroprotective in a Drosophila model of amyotrophic lateral sclerosis. J. Neurosci. 34, 15962–15974 (2014).

Download references


We thank K. R. Moss and K. T. Thomas for help with the preparation of primary cortical neurons, G. J. Bassell for logistical support and M. Castanedes-Casey for expert staining of human brain tissue. For numerous expression plasmids used in this study (Supplementary Table 3), we thank M. Hetzer (The Salk Institute for Biological Studies), J. Ellenberg (EMBL Heidelberg), V. Doye (Institut Jacques Monod, Université Paris Diderot/CNRS), J. Teodoro (McGill University) and J. Joseph (National Centre for Cell Science, S.P. Pune University), L. Gerace (The Scripps Research Institute), B. Paschal (University of Virginia School of Medicine) and M. Dasso (Eunice Kennedy Shriver National Institute of Child Health and Human Development). We thank the Bloomington Drosophila Stock Center for fly lines and Emory Integrated Proteomics Core, Neuropathology/ Histochemistry Core and Robert P. Apkarian Integrated Electron Microscopy Core for technical support. This work was supported by grants from the ALS Association (17-IIP-353) to W.R. and (16-IIP-278) to R.S.; the Emory Medicine Catalyst Funding Program to W.R.; Muscular Dystrophy Association (MDA348086) to R.S.; NIH grants K08-NS087121 to C.M.H., P30-NS055077 to the Neuropathology/Histochemistry core of the Emory NINDS Neurosciences Core Facility, AG025688 to Emory’s Alzheimer’s Disease Research Center, NIH R01-NS091299 to D.C.Z., R35-NS097261 to R.R., R01-NS085207 to R.S., R01NS091749 to W.R. and R01-NS093362 to W.R. and T.K., who is also supported by The Bluefield Project to Cure FTD; the Alzheimer’s Drug Discovery Foundation to N.J.C; and NIH R01-AG053960 to N.T.S., who is also supported in part by the Alzheimer’s Association (ALZ), Alzheimer’s Research UK (ARUK), The Michael J. Fox Foundation for Parkinson’s Research (MJFF) and a Weston Brain Institute Biomarkers Across Neurodegenerative Diseases Grant (11060). S.V. was partially funded by UBRP with funds from the UA Provost’s Office. P.G.D.-A. was funded by an ARCS Fellowship Roche Foundation Award.

Author information

Author notes

    • Ching-Chieh Chou

    Present address: Department of Biology, Stanford University, Stanford, CA, USA

    • Yi Zhang

    Present address: Department of Respiratory Medicine, The Second Xiangya Hospital, Central South University, Changsha, Hunan, China

    • Paul G. Donlin-Asp

    Present address: Max Planck Institute for Brain Research, Frankfurt, Germany

  1. Ching-Chieh Chou and Yi Zhang contributed equally to this work.


  1. Department of Cell Biology, Emory University School of Medicine, Atlanta, GA, USA

    • Ching-Chieh Chou
    • , Yi Zhang
    • , Paul G. Donlin-Asp
    • , Yu Han Chen
    • , Maureen A. Powers
    •  & Wilfried Rossoll
  2. Center for Neurodegenerative Disease, Emory University School of Medicine, Atlanta, GA, USA

    • Ching-Chieh Chou
    • , Mfon E. Umoh
    • , Duc M. Duong
    • , Nicholas T. Seyfried
    • , Thomas Kukar
    • , Chadwick M. Hales
    • , Marla Gearing
    • , Jonathan D. Glass
    •  & Wilfried Rossoll
  3. Xiangya Hospital and Xiangya School of Medicine, Central South University, Changsha, Hunan, China

    • Yi Zhang
  4. Department of Neurology, Emory University School of Medicine, Atlanta, GA, USA

    • Mfon E. Umoh
    • , Nicholas T. Seyfried
    • , Thomas Kukar
    • , Chadwick M. Hales
    • , Marla Gearing
    •  & Jonathan D. Glass
  5. Department of Molecular & Cellular Biology, University of Arizona, Tucson, AZ, USA

    • Spencer W. Vaughan
    • , Melissa Sayegh
    •  & Daniela C. Zarnescu
  6. Department of Neurobiology, Barrow Neurological Institute, Phoenix, AZ, USA

    • Ileana Lorenzini
    •  & Rita Sattler
  7. Department of Neuroscience, Mayo Clinic, Jacksonville, FL, USA

    • Feilin Liu
    • , Dennis W. Dickson
    • , Rosa Rademakers
    • , Yong-Jie Zhang
    • , Leonard Petrucelli
    •  & Wilfried Rossoll
  8. Department of Ophthalmology, the Second Hospital of Jilin University, Changchun, China

    • Feilin Liu
  9. Department of Biochemistry, Emory University School of Medicine, Atlanta, GA, USA

    • Duc M. Duong
    •  & Nicholas T. Seyfried
  10. Department of Pharmacology, Emory University School of Medicine, Atlanta, GA, USA

    • Thomas Kukar
  11. Department of Pathology and Laboratory Medicine, Emory University School of Medicine, Atlanta, GA, USA

    • Marla Gearing
  12. Department of Pathology and Immunology, Washington University, St. Louis, MO, USA

    • Nigel J. Cairns
  13. Department of Neurology, Mayo Clinic, Jacksonville, FL, USA

    • Kevin B. Boylan
  14. Emory ALS Center, Emory University School of Medicine, Atlanta, GA, USA

    • Jonathan D. Glass


  1. Search for Ching-Chieh Chou in:

  2. Search for Yi Zhang in:

  3. Search for Mfon E. Umoh in:

  4. Search for Spencer W. Vaughan in:

  5. Search for Ileana Lorenzini in:

  6. Search for Feilin Liu in:

  7. Search for Melissa Sayegh in:

  8. Search for Paul G. Donlin-Asp in:

  9. Search for Yu Han Chen in:

  10. Search for Duc M. Duong in:

  11. Search for Nicholas T. Seyfried in:

  12. Search for Maureen A. Powers in:

  13. Search for Thomas Kukar in:

  14. Search for Chadwick M. Hales in:

  15. Search for Marla Gearing in:

  16. Search for Nigel J. Cairns in:

  17. Search for Kevin B. Boylan in:

  18. Search for Dennis W. Dickson in:

  19. Search for Rosa Rademakers in:

  20. Search for Yong-Jie Zhang in:

  21. Search for Leonard Petrucelli in:

  22. Search for Rita Sattler in:

  23. Search for Daniela C. Zarnescu in:

  24. Search for Jonathan D. Glass in:

  25. Search for Wilfried Rossoll in:


W.R. conceived and directed the project. C.-C.C., Y.Z. and W.R. designed the experiments. C.-C.C. and W.R. interpreted data and wrote the manuscript. C.-C.C. characterized protein interactome, performed bioinformatic analysis and conducted experiments in N2a cells, primary cortical neurons and human fibroblasts with help from F.L. and Y.H.C. Y.Z. performed the BioID pulldown and sample preparation for LC-MS/MS analysis. M.E.U. performed immunohistochemistry staining. S.W.V., M.S. and D.C.Z. performed Drosophila experiments. I.L. and R.S. performed experiments in iPSC-derived motor neurons. C.-C.C. and P.G.D.-A. conducted SIM experiments. D.M.D., N.T.S. and M.A.P. provided technical support. T.K. provided key reagents. C.M.H., M.G., N.J.C., K.B.B., D.W.D., R.R., Y.-J.Z., L.P. and J.D.G. provided patient tissue with associated clinical and genetics data.

Competing interests

The authors declare that they have no competing financial interests. 

Corresponding author

Correspondence to Wilfried Rossoll.

Supplementary information

  1. Supplementary Text and Figures

    Supplementary Figures 1–17 and Supplementary Tables 1–4

  2. Life Sciences Reporting Summary

  3. Supplementary Video 1: GFP-expressing cells show normal NM morphology

    Three-dimensional (3D) reconstruction of nuclear membrane (NM) staining with anti-lamin B antibody (red) in primary neurons expressing GFP (green).

  4. Supplementary Video 2: GFP-CTF-expressing cells show abnormal NM morphology with deep invaginations

    Three-dimensional (3D) reconstruction of nuclear membrane (NM) staining with anti-lamin B antibody (red) in primary neurons expressing GFP-TDP-CTF (green).

  5. Supplementary Data

    Proteomics list

About this article

Publication history




Issue Date



Further reading