Abstract
The DNA- and RNA-binding protein fused in sarcoma (FUS) has been pathologically and genetically linked to amyotrophic lateral sclerosis (ALS) or frontotemporal lobar degeneration (FTLD). Cytoplasmic FUS-positive inclusions were identified in the brain and spinal cord of a subset of patients suffering with ALS/FTLD. An increasing number of reports suggest that FUS protein can behave in a prion-like manner. However, no neuropathological studies or experimental data were available regarding cell-to-cell spread of these pathological protein assemblies. In the present report, we investigated the ability of wild-type and mutant forms of FUS to transfer between neuronal cells. We combined the use of Drosophila models for FUS proteinopathies with that of the primary neuronal cultures to address neuron-to-neuron transfer of FUS proteins. Using conditional co-culture models and an optimized flow cytometry-based methodology, we demonstrated that ALS-mutant forms of FUS proteins can transfer between well-differentiated mature Drosophila neurons. These new observations support that a propagating mechanism could be applicable to FUS, leading to the sequential dissemination of pathological proteins over years.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Alliegro MC, Alliegro MA (1996) A nuclear protein regulated during the transition from active to quiescent phenotype in cultured endothelial cells. Dev Biol 174:288–297
Alonso AD, Beharry C, Corbo CP, Cohen LS (2016) Molecular mechanism of prion-like tau-induced neurodegeneration. Alzheimers Dement 12:1090–1097
Ashraf SI, McLoon AL, Sclarsic SM, Kunes S (2006) Synaptic protein synthesis associated with memory is regulated by the RISC pathway in Drosophila. Cell 124:191–205
Bosco DA, Lemay N, Ko HK, Zhou H, Burke C, Kwiatkowski TJ Jr, Sapp P, McKenna-Yasek D, Brown RH Jr, Hayward LJ (2010) Mutant FUS proteins that cause amyotrophic lateral sclerosis incorporate into stress granules. Hum Mol Genet 19:4160–4175
Casci I, Pandey UB (2015) A fruitful endeavor: modeling ALS in the fruit fly. Brain Res 1607:47–74
Chen Y, Yang M, Deng J, Chen X, Ye Y, Zhu L, Liu J, Ye H, Shen Y, Li Y et al (2011) Expression of human FUS protein in Drosophila leads to progressive neurodegeneration. Protein Cell 2:477–486
Chio A, Restagno G, Brunetti M, Ossola I, Calvo A, Mora G, Sabatelli M, Monsurro MR, Battistini S, Mandrioli J et al (2009) Two Italian kindreds with familial amyotrophic lateral sclerosis due to FUS mutation. Neurobiol Aging 30:1272–1275
Couthouis J, Hart MP, Shorter J, DeJesus-Hernandez M, Erion R, Oristano R, Liu AX, Ramos D, Jethava N, Hosangadi D et al (2011) A yeast functional screen predicts new candidate ALS disease genes. Proc Natl Acad Sci U S A 108:20881–20890
Cushman M, Johnson BS, King OD, Gitler AD, Shorter J (2010) Prion-like disorders: blurring the divide between transmissibility and infectivity. J Cell Sci 123:1191–1201
DeJesus-Hernandez M, Kocerha J, Finch N, Crook R, Baker M, Desaro P, Johnston A, Rutherford N, Wojtas A, Kennelly K et al (2010) De novo truncating FUS gene mutation as a cause of sporadic amyotrophic lateral sclerosis. Hum Mutat 31:E1377–E1389
Deng J, Yang M, Chen Y, Chen X, Liu J, Sun S, Cheng H, Li Y, Bigio EH, Mesulam M et al (2015) FUS interacts with HSP60 to promote mitochondrial damage. PLoS Genet 11:e1005357
Dormann D, Haass C (2011) TDP-43 and FUS: a nuclear affair. Trends Neurosci 34:339–348
Dormann D, Rodde R, Edbauer D, Bentmann E, Fischer I, Hruscha A, Than ME, Mackenzie IR, Capell A, Schmid B et al (2010) ALS-associated fused in sarcoma (FUS) mutations disrupt Transportin-mediated nuclear import. EMBO J 29:2841–2857
Fujii R, Takumi T (2005) TLS facilitates transport of mRNA encoding an actin-stabilizing protein to dendritic spines. J Cell Sci 118:5755–5765
Fujii R, Okabe S, Urushido T, Inoue K, Yoshimura A, Tachibana T, Nishikawa T, Hicks GG, Takumi T (2005) The RNA binding protein TLS is translocated to dendritic spines by mGluR5 activation and regulates spine morphology. Curr Biol 15:587–593
Hasegawa M (2016) Molecular mechanisms in the pathogenesis of Alzheimer’s disease and tauopathies-prion-like seeded aggregation and phosphorylation. Biomol Ther 6
Ju S, Tardiff DF, Han H, Divya K, Zhong Q, Maquat LE, Bosco DA, Hayward LJ, Brown RH Jr, Lindquist S et al (2011) A yeast model of FUS/TLS-dependent cytotoxicity. PLoS Biol 9:e1001052
Kamelgarn M, Chen J, Kuang L, Arenas A, Zhai J, Zhu H, Gal J (2016) Proteomic analysis of FUS interacting proteins provides insights into FUS function and its role in ALS. Biochim Biophys Acta 1862:2004–2014
Kino Y, Washizu C, Aquilanti E, Okuno M, Kurosawa M, Yamada M, Doi H, Nukina N (2011) Intracellular localization and splicing regulation of FUS/TLS are variably affected by amyotrophic lateral sclerosis-linked mutations. Nucleic Acids Res 39:2781–2798
Kwiatkowski TJ Jr, Bosco DA, Leclerc AL, Tamrazian E, Vanderburg CR, Russ C, Davis A, Gilchrist J, Kasarskis EJ, Munsat T et al (2009) Mutations in the FUS/TLS gene on chromosome 16 cause familial amyotrophic lateral sclerosis. Science 323:1205–1208
Lahiani-Skiba M, Barbot C, Bounoure F, Joudieh S, Skiba M (2006) Solubility and dissolution rate of progesterone-cyclodextrin-polymer systems. Drug Dev Ind Pharm 32:1043–1058
Lee S, Kim HJ (2015) Prion-like mechanism in amyotrophic lateral sclerosis: are protein aggregates the key? Exp Neurobiol 24:1–7
Meissner M, Lopato S, Gotzmann J, Sauermann G, Barta A (2003) Proto-oncoprotein TLS/FUS is associated to the nuclear matrix and complexed with splicing factors PTB, SRm160, and SR proteins. Exp Cell Res 283:184–195
Miguel L, Avequin T, Delarue M, Feuillette S, Frebourg T, Campion D, Lecourtois M (2012) Accumulation of insoluble forms of FUS protein correlates with toxicity in drosophila. Neurobiol Aging 33(1008):e1001–e1015
Nomura T, Watanabe S, Kaneko K, Yamanaka K, Nukina N, Furukawa Y (2014) Intranuclear aggregation of mutant FUS/TLS as a molecular pathomechanism of amyotrophic lateral sclerosis. J Biol Chem 289:1192–1202
Osterwalder T, Yoon KS, White BH, Keshishian H (2001) A conditional tissue-specific transgene expression system using inducible GAL4. Proc Natl Acad Sci U S A 98:12596–12601
Ratti A, Buratti E (2016) Physiological functions and pathobiology of TDP-43 and FUS/TLS proteins. J Neurochem 138(Suppl 1):95–111
Ravits J, Appel S, Baloh RH, Barohn R, Brooks BR, Elman L, Floeter MK, Henderson C, Lomen-Hoerth C, Macklis JD et al (2013) Deciphering amyotrophic lateral sclerosis: what phenotype, neuropathology and genetics are telling us about pathogenesis. Amyotroph Lateral Scler Frontotemporal Degener 14(Suppl 1):5–18
Rivero-Gutiérrez B, Anzola A, Martínez-Augustin O, de Medina FS (2014) Stain-free detection as loading control alternative to ponceau and housekeeping protein immunodetection in Western blotting. Anal Biochem 467:1–3
Schwartz JC, Podell ER, Han SSW, Berry JD, Eggan KC, Cech TR (2014) FUS is sequestered in nuclear aggregates in ALS patient fibroblasts. Mol Biol Cell 25:2571–2578
Smethurst P, Sidle KC, Hardy J (2015) Review: Prion-like mechanisms of transactive response DNA binding protein of 43 kDa (TDP-43) in amyotrophic lateral sclerosis (ALS). Neuropathol Appl Neurobiol 41:578–597
Sun Z, Diaz Z, Fang X, Hart MP, Chesi A, Shorter J, Gitler AD (2011) Molecular determinants and genetic modifiers of aggregation and toxicity for the ALS disease protein FUS/TLS. PLoS Biol 9:e1000614
Tateishi T, Hokonohara T, Yamasaki R, Miura S, Kikuchi H, Iwaki A, Tashiro H, Furuya H, Nagara Y, Ohyagi Y et al (2010) Multiple system degeneration with basophilic inclusions in Japanese ALS patients with FUS mutation. Acta Neuropathol 119:355–364
Tyson T, Steiner JA, Brundin P (2015) Sorting out release, uptake and processing of alpha-synuclein during prion-like spread of pathology. J Neurochem 139(Suppl 1):275–289
Urwin H, Josephs KA, Rohrer JD, Mackenzie IR, Neumann M, Authier A, Seelaar H, Van Swieten JC, Brown JM, Johannsen P et al (2010) FUS pathology defines the majority of tau- and TDP-43-negative frontotemporal lobar degeneration. Acta Neuropathol 120:33–41
Vance C, Rogelj B, Hortobagyi T, De Vos KJ, Nishimura AL, Sreedharan J, Hu X, Smith B, Ruddy D, Wright P et al (2009) Mutations in FUS, an RNA processing protein, cause familial amyotrophic lateral sclerosis type 6. Science 323:1208–1211
Venken KJ, Simpson JH, Bellen HJ (2011) Genetic manipulation of genes and cells in the nervous system of the fruit fly. Neuron 72:202–230
Walker LC, Schelle J, Jucker M (2016) The prion-like properties of amyloid-beta assemblies: implications for Alzheimer’s disease. Cold Spring Harb Perspect Med 6
Acknowledgements
We thank the Bloomington Drosophila stock center for providing fly stocks. Confocal images have been obtained at the PRIMACEN imaging platform (Rouen University, France). Flow cytometry and FACS experiments were performed at the CyFlow platform (Rouen University, France). We thank Damien Schapman and Sahil Adriouch for their technical assistance and helpful discussions. This work was co-supported by a grant from the France Alzheimer Association to ML, the European Union, and the Région Normandie. Europe gets involved in Normandie with European Regional Development Fund (ERDF). MD is a PhD fellow of the Région Normandie.
Author information
Authors and Affiliations
Contributions
S.F., M.D., G. R., and A-L. G. performed the experiments and analyzed the results. M.L., D.C, J.W., and T.F supervised the project. M. L. wrote the manuscript with M.D., O.B., and S.F. Funding were obtained by M.L., D.C., T.F., O.B., and Z.L.
Corresponding author
Ethics declarations
Competing Interests
The authors declare that they have no competing interests.
Electronic supplementary material
ESM 1
(PDF 5808 kb)
Rights and permissions
About this article
Cite this article
Feuillette, S., Delarue, M., Riou, G. et al. Neuron-to-Neuron Transfer of FUS in Drosophila Primary Neuronal Culture Is Enhanced by ALS-Associated Mutations. J Mol Neurosci 62, 114–122 (2017). https://doi.org/10.1007/s12031-017-0908-y
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s12031-017-0908-y