A role of cellular translation regulation associated with toxic Huntingtin protein

Abstract

Huntington’s disease (HD) is a severe neurodegenerative disorder caused by poly Q repeat expansion in the Huntingtin (Htt) gene. While the Htt amyloid aggregates are known to affect many cellular processes, their role in translation has not been addressed. Here we report that pathogenic Htt expression causes a protein synthesis deficit in cells. We find a functional prion-like protein, the translation regulator Orb2, to be sequestered by Htt aggregates in cells. Co-expression of Orb2 can partially rescue the lethality associated with poly Q expanded Htt. These findings can be relevant for HD as human homologs of Orb2 are also sequestered by pathogenic Htt aggregates. Our work suggests that translation dysfunction is one of the contributors to the pathogenesis of HD and new therapies targeting protein synthesis pathways might help to alleviate disease symptoms.

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References

  1. 1.

    Bates GP, Dorsey R, Gusella JF et al (2015) Huntington disease. Nat Rev Dis Primers 1:15005. https://doi.org/10.1038/nrdp.2015.5

    Article  PubMed  Google Scholar 

  2. 2.

    Orr HT, Zoghbi HY (2007) Trinucleotide repeat disorders. Annu Rev Neurosci 30:575–621. https://doi.org/10.1146/annurev.neuro.29.051605.113042

    CAS  Article  PubMed  Google Scholar 

  3. 3.

    MacDonald ME, Ambrose CM, Duyao MP et al (1993) A novel gene containing a trinucleotide repeat that is expanded and unstable on Huntington’s disease chromosomes. Cell 72:971–983. https://doi.org/10.1016/0092-8674(93)90585-E

    Article  Google Scholar 

  4. 4.

    Lee J-M, Ramos EM, Lee J-H et al (2012) CAG repeat expansion in Huntington disease determines age at onset in a fully dominant fashion. Neurology 78:690–695. https://doi.org/10.1212/WNL.0b013e318249f683

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  5. 5.

    Klaips CL, Jayaraj GG, Hartl FU (2018) Pathways of cellular proteostasis in aging and disease. J Cell Biol 217:51–63. https://doi.org/10.1083/jcb.201709072

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Bennett EJ, Shaler TA, Woodman B et al (2007) Global changes to the ubiquitin system in Huntington’s disease. Nature 448:704–708. https://doi.org/10.1038/nature06022

    CAS  Article  PubMed  Google Scholar 

  7. 7.

    Hageman J, Rujano MA, van Waarde MAWH et al (2010) A DNAJB chaperone subfamily with HDAC-dependent activities suppresses toxic protein aggregation. Mol Cell 37:355–369. https://doi.org/10.1016/j.molcel.2010.01.001

    CAS  Article  PubMed  Google Scholar 

  8. 8.

    Hay DG, Sathasivam K, Tobaben S et al (2004) Progressive decrease in chaperone protein levels in a mouse model of Huntington’s disease and induction of stress proteins as a therapeutic approach. Hum Mol Genet 13:1389–1405. https://doi.org/10.1093/hmg/ddh144

    CAS  Article  PubMed  Google Scholar 

  9. 9.

    Hipp MS, Patel CN, Bersuker K et al (2012) Indirect inhibition of 26S proteasome activity in a cellular model of Huntington’s disease. J Cell Biol 196:573–587. https://doi.org/10.1083/jcb.201110093

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  10. 10.

    Labbadia J, Novoselov SS, Bett JS et al (2012) Suppression of protein aggregation by chaperone modification of high molecular weight complexes. Brain 135:1180–1196. https://doi.org/10.1093/brain/aws022

    Article  PubMed  PubMed Central  Google Scholar 

  11. 11.

    Martinez-Vicente M, Talloczy Z, Wong E et al (2010) Cargo recognition failure is responsible for inefficient autophagy in Huntington’s disease. Nat Neurosci 13:567–576. https://doi.org/10.1038/nn.2528

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  12. 12.

    Reis SD, Pinho BR, Oliveira JMA (2017) Modulation of molecular chaperones in Huntington’s disease and other polyglutamine disorders. Mol Neurobiol 54:5829–5854. https://doi.org/10.1007/s12035-016-0120-z

    CAS  Article  PubMed  Google Scholar 

  13. 13.

    Tauber E, Miller-Fleming L, Mason RP et al (2011) Functional gene expression profiling in yeast implicates translational dysfunction in mutant huntingtin toxicity. J Biol Chem 286:410–419. https://doi.org/10.1074/jbc.M110.101527

    CAS  Article  PubMed  Google Scholar 

  14. 14.

    Yang J, Hao X, Cao X et al (2016) Spatial sequestration and detoxification of Huntingtin by the ribosome quality control complex. Elife. https://doi.org/10.7554/eLife.11792

    Article  PubMed  PubMed Central  Google Scholar 

  15. 15.

    Culver BP, Savas JN, Park SK et al (2012) Proteomic analysis of wild-type and mutant huntingtin-associated proteins in mouse brains identifies unique interactions and involvement in protein synthesis. J Biol Chem 287:21599–21614. https://doi.org/10.1074/jbc.M112.359307

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Kim YE, Hosp F, Frottin F et al (2016) Soluble Oligomers of PolyQ-expanded huntingtin target a multiplicity of key cellular factors. Mol Cell 63:951–964. https://doi.org/10.1016/j.molcel.2016.07.022

    CAS  Article  PubMed  Google Scholar 

  17. 17.

    Hosp F, Gutiérrez-Ángel S, Schaefer MH et al (2017) Spatiotemporal proteomic profiling of Huntington’s disease inclusions reveals widespread loss of protein function. Cell Rep 21:2291–2303. https://doi.org/10.1016/j.celrep.2017.10.097

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Bañez-Coronel M, Ayhan F, Tarabochia AD et al (2015) RAN translation in huntington disease. Neuron 88:667–677. https://doi.org/10.1016/j.neuron.2015.10.038

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  19. 19.

    Graham RK, Deng Y, Slow EJ et al (2006) Cleavage at the caspase-6 site is required for neuronal dysfunction and degeneration due to mutant huntingtin. Cell 125:1179–1191. https://doi.org/10.1016/j.cell.2006.04.026

    CAS  Article  PubMed  Google Scholar 

  20. 20.

    Warner JR, Knopf PM, Rich A (1963) A multiple ribosomal structure in protein synthesis. Proc Natl Acad Sci USA 49:122–129. https://doi.org/10.1073/pnas.49.1.122

    CAS  Article  PubMed  Google Scholar 

  21. 21.

    Meriin AB, Zhang X, He X et al (2002) Huntingtin toxicity in yeast model depends on polyglutamine aggregation mediated by a prion-like protein Rnq1. J Cell Biol 157:997–1004. https://doi.org/10.1083/jcb.200112104

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Nathans D (1964) Puromycin inhibition of protein synthesis: incorporation of puromycin into peptide chains. PNAS 51:585–592. https://doi.org/10.1073/pnas.51.4.585

    CAS  Article  PubMed  Google Scholar 

  23. 23.

    Deliu LP, Ghosh A, Grewal SS (2017) Investigation of protein synthesis in Drosophila larvae using puromycin labelling. Biol Open 6:1229–1234. https://doi.org/10.1242/bio.026294

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Schmidt EK, Clavarino G, Ceppi M, Pierre P (2009) SUnSET, a nonradioactive method to monitor protein synthesis. Nat Methods 6:275–277. https://doi.org/10.1038/nmeth.1314

    CAS  Article  PubMed  Google Scholar 

  25. 25.

    Liu J, Xu Y, Stoleru D, Salic A (2012) Imaging protein synthesis in cells and tissues with an alkyne analog of puromycin. Proc Natl Acad Sci USA 109:413–418. https://doi.org/10.1073/pnas.1111561108

    Article  PubMed  Google Scholar 

  26. 26.

    Culver BP, DeClercq J, Dolgalev I et al (2016) Huntington’s disease protein Huntingtin associates with its own mRNA. J Huntingtons Dis 5:39–51. https://doi.org/10.3233/JHD-150177

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  27. 27.

    Hervás R, Li L, Majumdar A et al (2016) Molecular Basis of Orb2 amyloidogenesis and blockade of memory consolidation. PLoS Biol 14:e1002361. https://doi.org/10.1371/journal.pbio.1002361

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  28. 28.

    Ivshina M, Lasko P, Richter JD (2014) Cytoplasmic polyadenylation element binding proteins in development, health, and disease. Annu Rev Cell Dev Biol 30:393–415. https://doi.org/10.1146/annurev-cellbio-101011-155831

    CAS  Article  PubMed  Google Scholar 

  29. 29.

    Keleman K, Krüttner S, Alenius M, Dickson BJ (2007) Function of the Drosophila CPEB protein Orb2 in long-term courtship memory. Nat Neurosci 10:1587–1593. https://doi.org/10.1038/nn1996

    CAS  Article  PubMed  Google Scholar 

  30. 30.

    Majumdar A, Cesario WC, White-Grindley E et al (2012) Critical role of amyloid-like oligomers of Drosophila Orb2 in the persistence of memory. Cell 148:515–529. https://doi.org/10.1016/j.cell.2012.01.004

    CAS  Article  PubMed  Google Scholar 

  31. 31.

    Khan MR, Li L, Pérez-Sánchez C et al (2015) Amyloidogenic oligomerization transforms Drosophila Orb2 from a translation repressor to an activator. Cell 163:1468–1483. https://doi.org/10.1016/j.cell.2015.11.020

    CAS  Article  PubMed  Google Scholar 

  32. 32.

    Screening for Amyloid Aggregation by Semi-Denaturing Detergent-Agarose Gel Electrophoresis| Protocol. https://www.jove.com/video/838/screening-for-amyloid-aggregation-semi-denaturing-detergent-agarose. Accessed 28 Oct 2019

  33. 33.

    Ding Q, Markesbery WR, Chen Q et al (2005) Ribosome dysfunction is an early event in Alzheimer’s disease. J Neurosci 25:9171–9175. https://doi.org/10.1523/JNEUROSCI.3040-05.2005

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  34. 34.

    Russo A, Scardigli R, La Regina F et al (2017) Increased cytoplasmic TDP-43 reduces global protein synthesis by interacting with RACK1 on polyribosomes. Hum Mol Genet 26:1407–1418. https://doi.org/10.1093/hmg/ddx035

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  35. 35.

    López-Erauskin J, Tadokoro T, Baughn MW et al (2018) ALS/FTD-linked mutation in FUS suppresses intra-axonal protein synthesis and drives disease without nuclear loss-of-function of FUS. Neuron 100:816–830.e7. https://doi.org/10.1016/j.neuron.2018.09.044

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  36. 36.

    Kamelgarn M, Chen J, Kuang L et al (2018) ALS mutations of FUS suppress protein translation and disrupt the regulation of nonsense-mediated decay. Proc Natl Acad Sci USA 115:E11904–E11913. https://doi.org/10.1073/pnas.1810413115

    CAS  Article  PubMed  Google Scholar 

  37. 37.

    Hartmann H, Hornburg D, Czuppa M, et al (2018) Proteomics and C9orf72 neuropathology identify ribosomes as poly-GR/PR interactors driving toxicity. Life Sci Alliance. https://doi.org/10.26508/lsa.201800070

  38. 38.

    Zhang Y-J, Gendron TF, Ebbert MTW et al (2018) Poly(GR) impairs protein translation and stress granule dynamics in C9orf72 -associated frontotemporal dementia and amyotrophic lateral sclerosis. Nat Med 24:1136. https://doi.org/10.1038/s41591-018-0071-1

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  39. 39.

    Kanekura K, Yagi T, Cammack AJ et al (2016) Poly-dipeptides encoded by the C9ORF72 repeats block global protein translation. Hum Mol Genet 25:1803–1813. https://doi.org/10.1093/hmg/ddw052

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Moens TG, Niccoli T, Wilson KM et al (2019) C9orf72 arginine-rich dipeptide proteins interact with ribosomal proteins in vivo to induce a toxic translational arrest that is rescued by eIF1A. Acta Neuropathol 137:487–500. https://doi.org/10.1007/s00401-018-1946-4

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  41. 41.

    Meier S, Bell M, Lyons DN et al (2016) Pathological tau promotes neuronal damage by impairing ribosomal function and decreasing protein synthesis. J Neurosci 36:1001–1007. https://doi.org/10.1523/JNEUROSCI.3029-15.2016

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  42. 42.

    Eshraghi M, Karunadharma P, Blin J et al (2019) Global ribosome profiling reveals that mutant huntingtin stalls ribosomes and represses protein synthesis independent of fragile X mental retardation protein. https://doi.org/10.1101/629667

  43. 43.

    Furukawa Y, Kaneko K, Matsumoto G et al (2009) Cross-seeding fibrillation of Q/N-rich proteins offers new pathomechanism of polyglutamine diseases. J Neurosci 29:5153–5162. https://doi.org/10.1523/JNEUROSCI.0783-09.2009

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  44. 44.

    Giasson BI, Forman MS, Higuchi M et al (2003) Initiation and synergistic fibrillization of tau and alpha-synuclein. Science 300:636–640. https://doi.org/10.1126/science.1082324

    CAS  Article  PubMed  Google Scholar 

  45. 45.

    Guo JL, Covell DJ, Daniels JP et al (2013) Distinct α-synuclein strains differentially promote tau inclusions in neurons. Cell 154:103–117. https://doi.org/10.1016/j.cell.2013.05.057

    CAS  Article  PubMed  Google Scholar 

  46. 46.

    Katorcha E, Makarava N, Lee YJ et al (2017) Cross-seeding of prions by aggregated α-synuclein leads to transmissible spongiform encephalopathy. PLoS Pathog 13:e1006563. https://doi.org/10.1371/journal.ppat.1006563

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  47. 47.

    Ono K, Takahashi R, Ikeda T, Yamada M (2012) Cross-seeding effects of amyloid β-protein and α-synuclein. J Neurochem 122:883–890. https://doi.org/10.1111/j.1471-4159.2012.07847.x

    CAS  Article  PubMed  Google Scholar 

  48. 48.

    Tanaka M, Ishizuka K, Nekooki-Machida Y et al (2017) Aggregation of scaffolding protein DISC1 dysregulates phosphodiesterase 4 in Huntington’s disease. J Clin Invest 127:1438–1450. https://doi.org/10.1172/JCI85594

    Article  PubMed  PubMed Central  Google Scholar 

  49. 49.

    Bilen J, Bonini NM (2007) Genome-wide screen for modifiers of Ataxin-3 neurodegeneration in Drosophila. PLoS Genet 3:e177. https://doi.org/10.1371/journal.pgen.0030177

    CAS  Article  PubMed Central  Google Scholar 

  50. 50.

    Burguete AS, Almeida S, Gao F-B et al (2015) GGGGCC microsatellite RNA is neuritically localized, induces branching defects, and perturbs transport granule function. eLife 4:e08881. https://doi.org/10.7554/eLife.08881

  51. 51.

    Ripaud L, Chumakova V, Antonin M et al (2014) Overexpression of Q-rich prion-like proteins suppresses polyQ cytotoxicity and alters the polyQ interactome. Proc Natl Acad Sci USA 111:18219–18224. https://doi.org/10.1073/pnas.1421313111

    CAS  Article  PubMed  Google Scholar 

  52. 52.

    Kayatekin C, Matlack KES, Hesse WR et al (2014) Prion-like proteins sequester and suppress the toxicity of huntingtin exon 1. Proc Natl Acad Sci USA 111:12085–12090. https://doi.org/10.1073/pnas.1412504111

    CAS  Article  PubMed  Google Scholar 

  53. 53.

    Stepien BK, Oppitz C, Gerlach D et al (2016) RNA-binding profiles of Drosophila CPEB proteins Orb and Orb2. PNAS 113:E7030–E7038. https://doi.org/10.1073/pnas.1603715113

    CAS  Article  PubMed  Google Scholar 

  54. 54.

    Fayazi Z, Ghosh S, Marion S et al (2006) A Drosophila ortholog of the human MRJ modulates polyglutamine toxicity and aggregation. Neurobiol Dis 24:226–244. https://doi.org/10.1016/j.nbd.2006.06.015

    CAS  Article  PubMed  Google Scholar 

  55. 55.

    Kazemi-Esfarjani P, Benzer S (2000) Genetic suppression of polyglutamine toxicity in Drosophila. Science 287:1837–1840. https://doi.org/10.1126/science.287.5459.1837

    CAS  Article  PubMed  Google Scholar 

  56. 56.

    Belloc E, Piqué M, Méndez R (2008) Sequential waves of polyadenylation and deadenylation define a translation circuit that drives meiotic progression. Biochem Soc Trans 36:665–670. https://doi.org/10.1042/BST0360665

    CAS  Article  PubMed  Google Scholar 

  57. 57.

    Piqué M, López JM, Foissac S et al (2008) A combinatorial code for CPE-mediated translational control. Cell 132:434–448. https://doi.org/10.1016/j.cell.2007.12.038

    CAS  Article  PubMed  Google Scholar 

  58. 58.

    Parras A, Anta H, Santos-Galindo M et al (2018) Autism-like phenotype and risk gene mRNA deadenylation by CPEB4 mis-splicing. Nature 560:441–446. https://doi.org/10.1038/s41586-018-0423-5

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  59. 59.

    Aldaz T, Nigro P, Sánchez-Gómez A et al (2019) Non-motor symptoms in Huntington’s disease: a comparative study with Parkinson’s disease. J Neurol 266:1340–1350. https://doi.org/10.1007/s00415-019-09263-7

    Article  PubMed  Google Scholar 

  60. 60.

    Carmichael AM, Irish M, Glikmann-Johnston Y et al (2019) Pervasive autobiographical memory impairments in Huntington’s disease. Neuropsychologia 127:123–130. https://doi.org/10.1016/j.neuropsychologia.2019.02.017

    Article  PubMed  Google Scholar 

  61. 61.

    Weiss KR, Kimura Y, Lee W-CM, Littleton JT (2012) Huntingtin aggregation kinetics and their pathological role in a Drosophila Huntington’s disease model. Genetics 190:581–600. https://doi.org/10.1534/genetics.111.133710

    CAS  Article  PubMed  PubMed Central  Google Scholar 

  62. 62.

    Koulouras G, Panagopoulos A, Rapsomaniki MA et al (2018) EasyFRAP-web: a web-based tool for the analysis of fluorescence recovery after photobleaching data. Nucl Acids Res 46:W467–W472. https://doi.org/10.1093/nar/gky508

    CAS  Article  PubMed  Google Scholar 

  63. 63.

    Rapsomaniki MA, Kotsantis P, Symeonidou I-E et al (2012) easyFRAP: an interactive, easy-to-use tool for qualitative and quantitative analysis of FRAP data. Bioinformatics 28:1800–1801. https://doi.org/10.1093/bioinformatics/bts241

    CAS  Article  PubMed  Google Scholar 

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Acknowledgements

We acknowledge Prof. Troy Littleton, Prof. Kausik Si for sharing several Drosophila stocks and plasmids with us. We also thank Addgene and Bloomington Drosophila stock center for some of the plasmids and fly lines used in this work. We thank Prof. Kausik Si, Dr. Gunther Hollopeter, Dr. Irina Dudanova and Dr. Deepa Subramanyam for their suggestions on the work. We thank Dr. Vidisha Tripathi for letting us use the qPCR machine. A.M acknowledges Dr. Arvind Sahu, Dr. Vasudevan Seshadri and Dr. Ajay Pillai for their support. This work was supported by funding from Wellcome Trust-DBT India Alliance intermediate fellowship (IA/I/13/2/501030) to AM along with intramural funding from NCCS. TB was supported by Ramalingaswami fellowship from DBT (BT/RLF/Re-entry/54/2013) and IYBA Grant (BT/09/IYBA/2015/03).

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HJ started this project and identified Orb2 isoforms as rescuers of Htt pathogenicity. VG performed the S2 cell polysome and puromycin incorporation experiments. MS1 performed S2 cell imaging and FRAP experiments. MS2 did the Yeast polysome experiments. AR did the Orb2 rescue and Orb2 RNAi experiments. MD performed the Orb2 level quantitation, OPP staining, Htt clonings, and immunoprecipitation experiments. RC and AD performed the Yeast lethality and growth curve experiments. TB and AM conceived and designed the experiments. AM wrote the manuscript.

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Correspondence to Tania Bose or Amitabha Majumdar.

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(A)

FACS analysis of S2 cells transfected with HttQ15 and HttQ138 stained with 7AAD to check cell viability. No significant difference was observed with respect to 7AAD negative and RFP positive cells between HttQ15 and HttQ138 samples. Data is from 3 independent experiments and is represented as % live cells. Error bars represent SEM and significance is tested using unpaired one-tailed t-test (B) Cell counting of Trypan blue stained Yeast cells at 8 hours post galactose induction show no significant difference between cells expressing HttQ25 and HttQ103 (C) Yeast growth curve of HttQ25 and HttQ103 done with 3 independent repeats. At the 8 hour timepoint when polysome experiments were done the difference in growth in Q25 and Q103 is not significant (unpaired one-tailed t-test) with a p-value of 0.1363 (D) Quantitative real-time PCR to detect mRNA levels in Drosophila brains expressing HttQ15 and HttQ138 show no significant difference in endogenous transcript levels of Orb2. Data is from n=4 independent experiments and is represented as relative fold change in Orb2 levels for HttQ138 compared to HttQ15. Error bars represent SEM and significance is tested using unpaired one-tailed t-test and the p-value is 0.1661(ns) (E) Images of Neuro2A cells expressing RFP tagged HttQ15 and HttQ138. HttQ15 shows a diffused expression pattern while HttQ138 forms aggregate in these cell lines. 1 (PDF 2251 kb)

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Joag, H., Ghatpande, V., Desai, M. et al. A role of cellular translation regulation associated with toxic Huntingtin protein. Cell. Mol. Life Sci. 77, 3657–3670 (2020). https://doi.org/10.1007/s00018-019-03392-y

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Keywords

  • Huntington’s disease
  • Functional-prion-like protein
  • Translation regulator
  • Orb2