Skip to main content

Advertisement

Springer Nature Link
Log in

Bortezomib Induces the Formation of Nuclear poly(A) RNA Granules Enriched in Sam68 and PABPN1 in Sensory Ganglia Neurons

  • Published:
Neurotoxicity Research Aims and scope Submit manuscript

Abstract

The ubiquitin-dependent proteasome system (UPS) is the major pathway responsible for selective nuclear and cytoplasmic protein degradation. Bortezomib, a boronic acid dipeptide, is a reversible 20S proteasome inhibitor used as novel anticancer drug, particularly in the treatment of multiple myeloma and certain lymphomas. Bortezomib-induced peripheral neuropathy (BIPN) is a widely recognized dose-limiting neurotoxicity of this proteasome inhibitor, which causes a significant negative impact on the quality of life. The pathogenic mechanisms underlying bortezomib neurotoxicity are little known. In this study a rat was used as our animal model to investigate the bortezomib-induced nuclear changes in dorsal root ganglia (DRG) neurons. Our results indicate that this neuronal population is an important target of bortezomib neurotoxicity. Nuclear changes include accumulation of ubiquitin–protein conjugates, reduction of transcriptional activity, and nuclear retention of poly(A) RNAs in numerous spherical or ring-shaped dense granules. They also contained the RNA-binding proteins PABPN1 (poly(A) binding protein nuclear 1) and Sam68, but lacked the mRNA nuclear export factors REF and Y14. At the cytoplasmic level, most neurons exhibited chromatolysis, supporting the inhibition of mRNA translation. Our results indicate that bortezomib interferes with transcription, nuclear processing and transport, and cytoplasmic translation of mRNAs in DRG neurons. They also support that this neuronal dysfunction is an essential pathogenic mechanism in the BIPN, which is characterized by sensory impairment including sensory ataxia.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Price includes VAT (Canada)

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6

Similar content being viewed by others

References

  • Adams J (2004) The development of proteasome inhibitors as anticancer drugs. Cancer Cell 5:417–421

    Article  CAS  PubMed  Google Scholar 

  • Alves-Rodrigues A, Gregory L, Figueiredo-Pereira ME (1998) Ubiquitin, cellular inclusions and their role in neurodegeneration. Trends Neurosci 21:517–520

    Article  Google Scholar 

  • Anderson P, Kedersha N (2006) RNA granules. J Cell Biol 172:803–808

    Article  CAS  PubMed  Google Scholar 

  • Berciano MT, Villagra NT, Ojeda JL, Navascues J, Gomes A, Lafarga M, Carmo-Fonseca M (2004) Oculopharyngeal muscular dystrophy-like nuclear inclusions are present in normal magnocellular neurosecretory neurons of the hypothalamus. Hum Mol Genet 13:829–838

    Article  CAS  PubMed  Google Scholar 

  • Berciano MT, Novell M, Villagra NT, Casafont I, Bengoechea R, Val-Bernal JF, Lafarga M (2007) Cajal body number and nucleolar size correlate with the cell body mass in human sensory ganglia neurons. J Struct Biol 158:410–420

    Article  CAS  PubMed  Google Scholar 

  • Biamonti G (2004) Nuclear stress bodies: a heterochromatin affair? Nat Rev Mol Cell Biol 5:493–498

    Article  CAS  PubMed  Google Scholar 

  • Biggiogera M, Fakan S (1998) Fine structural specific visualization of RNA on ultrathin sections. J Histochem Cytochem 46:389–395

    CAS  PubMed  Google Scholar 

  • Biggiogera M, Cisterna B, Spedito A, Vecchio L, Malatesta M (2008) Perichromatin fibrils as early markers of transcriptional alterations. Differentiation 76:57–65

    CAS  PubMed  Google Scholar 

  • Bigotte L, Arvidson B, Olsson Y (1982) Cytofluorescence localization of adriamycin in the nervous system. II. Distribution of the drug in the somatic and autonomic peripheral nervous systems of normal adult mice after intravenous injection. Acta Neuropathol 57:130–136

    Article  CAS  PubMed  Google Scholar 

  • Carman GG, Mitchell HH (1926) Estimation of the surface area of the white rat. Am J Physiol 76:380–384

    Google Scholar 

  • Carter KC, Taneja KL, Lawrence JB (1991) Discrete nuclear domains of poly(A) RNA and their relationship to the functional organization of the nucleus. J Cell Biol 115:1191–1202

    Article  CAS  PubMed  Google Scholar 

  • Casafont I, Navascues J, Pena E, Lafarga M, Berciano MT (2006) Nuclear organization and dynamics of transcription sites in rat sensory ganglia neurons detected by incorporation of 5′-fluorouridina into nascent RNA. Neuroscience 140:453–462

    Article  CAS  PubMed  Google Scholar 

  • Cavaletti G, Gilardini A, Canta A, Rigamonti L, Rodriguez-Menendez V, Ceresa C, Marmiroli P, Bossi M, Oggioni N, D′Incalci M, De Coster R (2007) Bortezomib-induced peripheral neurotoxicity: a neurophysiological and pathological study in the rat. Exp Neurol 204:317–325

    Article  CAS  PubMed  Google Scholar 

  • Chen T, Boisvert FM, Bazzet-Jones DP, Richard S (1999) A role for the GSG domain in localizing Sam68 to novel nuclear structures in cancer cells. Mol Biol Cell 10:3015–3033

    CAS  PubMed  Google Scholar 

  • Chiodi I, Biggiogera M, Denegri M, Corioni M, Weighardt F, Cobianchi F, Riva S, Biamonti G (2000) Structure and dynamics of hnRNP-labelled nuclear bodies induced by stress treatments. J Cell Sci 113:4043–4053

    CAS  PubMed  Google Scholar 

  • Ciechanover A, DiGiuseppe JA, Bercovich B, Orian A, Richter JD (1991) Degradation of nuclear oncoproteins by the ubiquitin system in vitro. Proc Natl Acad Sci USA 88:139–143

    Article  CAS  PubMed  Google Scholar 

  • Cioce M, Lamond AI (2005) Cajal Bodies: A long history of discovery. Annu Rev Cell Dev Biol 21:105–131

    Article  CAS  PubMed  Google Scholar 

  • Collins GA, Tansey WP (2006) The proteasome: a utility tool for transcription? Curr Opin Genet Dev 16:197–202

    Article  CAS  PubMed  Google Scholar 

  • Dreyfuss G, Kim VN, Kataoka N (2002) Messenger-RNA-binding proteins and the messages they carry. Nat Rev Mol Cell Biol 3:195–205

    Article  CAS  PubMed  Google Scholar 

  • Dubois E (1936) Basal metabolism in health and disease. Lea & Febiger, Philadelphia, pp 125–144

    Google Scholar 

  • Filosto M, Rossi G, Pelizzari AM, Buzio S, Tentorio M, Broglio L, Mancuso M, Rinaldi M, Scarpelli M, Padovani A (2007) A high-dose bortezomib neuropathy with sensory ataxia and myelin involvement. J Neurol Sci 263:40–43

    Article  CAS  PubMed  Google Scholar 

  • Fredj NB, Grange J, Sadoul R, Richard S, Goldberg Y, Boyer V (2004) Depolarization-induced translocation of the RNA-binding protein Sam68 to the dendrites of hippocampal neurons. J Cell Sci 117:1079–1090

    Article  CAS  PubMed  Google Scholar 

  • Gall JC (2000) Cajal bodies: the first 100 years. Annu Rev Cell Dev Biol 16:273–300

    Article  CAS  PubMed  Google Scholar 

  • Grange J, Boyer V, Fabian-Fine R, Fredj NB, Sadoul R, Goldberg Y (2004) Somatodendritic localization and mRNA association of the splicing regulatory protein Sam68 in the hippocampus and cortex. J Neurosci Res 75:654–666

    Article  CAS  PubMed  Google Scholar 

  • Hartmann AM, Nayler O, Schwaiger FW, Obermeier A, Stamm S (1999) The interaction and colocalization of Sam68 with the splicing-associated factor YT521-B in nuclear dots is regulated by the Src family kinase p59fyn. Mol Biol Cell 10:3909–3926

    CAS  PubMed  Google Scholar 

  • Henao-Mejia J, Liu Y, Park YW, Zhang J, Sanford J, He JJ (2009) Supression of HIV-1 nef translation by Sam68 mutant-induced stress granules and nef mRNA sequestration. Mol Cell 33:87–96

    Article  CAS  PubMed  Google Scholar 

  • Jagannath S, Barlogie B, Berenson J, Siegel D, Irwin D, Richardson PG, Niesvizky R, Alexanian R, Limentani SA, Alsina M, Adams J, Kauffman M, Esseltine DL, Schenkein D, Anderson KC (2004) A phase 2 study of two doses of bortezomib in relapsed or refractory myeloma. Br J Haematol 127:165–172

    Article  CAS  PubMed  Google Scholar 

  • Krause S, Fakan S, Weis K, Wahle E (1994) Immunodetection of poly (A)-binding protein II in the cell nucleus. Exp Cell Res 214:75–82

    Article  CAS  PubMed  Google Scholar 

  • Kühn U, Wahle E (2004) Structure and function of poly(A) binding proteins. Biochim Biophys Acta 1678:67–84

    PubMed  Google Scholar 

  • Lafarga M, Berciano MT, Pena E, Mayo I, Castano JG, Bohmann D, Rodrigues JP, Tavanez JP, Carmo-Fonseca M (2002) Clastosome: a subtype of nuclear body enriched in 19S and 20S proteasomes, ubiquitin, and protein substrates of proteasome. Mol Biol Cell 13:2771–2782

    Article  CAS  PubMed  Google Scholar 

  • Lamond AI, Spector DL (2003) Nuclear speckles: a model for nuclear organelles. Nat Rev Mol Cell Biol 4:605–612

    Article  CAS  PubMed  Google Scholar 

  • Landowski TH, Megli CJ, Nullmeyer KD, Lynch RM, Dorr RT (2005) Mitochondrial-mediated disregulation of Ca2+ is a critical determinant of Velcade (PS-341/bortezomib) cytotoxicity in myeloma cells lines. Cancer Res 65:3828–3836

    Article  CAS  PubMed  Google Scholar 

  • Laroia G, Cuesta R, Brewer G, Scheneider RJ (1999) Control of mRNA decay by heat shock-ubiquitin-proteasome pathway. Science 284:499–502

    Article  CAS  PubMed  Google Scholar 

  • Laroia G, Sarkar B, Scheneider RJ (2002) Ubiquitin-dependent mechanisms regulates rapid turnover of AU-rich cytokine mRNAs. Proc Natl Acad Sci USA 99:1842–1846

    Article  CAS  PubMed  Google Scholar 

  • Li J, Liu Y, Kim BO, He JJ (2002) Direct participation of Sam68, the 68-kilodalton Src-associated protein in mitosis, in the CRM1-mediated Rev nuclear export pathway. J Virol 76:8374–8382

    Article  CAS  PubMed  Google Scholar 

  • Lukong KE, Richard S (2003) Sam68, the KH domain-containing superSTAR. Biochim Biophys Acta 1653:73–86

    CAS  PubMed  Google Scholar 

  • Matter N, Herrlich P, König H (2002) Signal-dependent regulation of splicing via phosphorylation of SAM68. Nature 420:691–695

    Article  CAS  PubMed  Google Scholar 

  • Mazroui R, Di Marco S, Kaufman RJ, Gallouzi IE (2007) Inhibition of the ubiquitin-proteasome system induces stress granule formation. Mol Biol Cell 18:2603–2618

    Article  CAS  PubMed  Google Scholar 

  • McConkey DJ, Zhu K (2008) Mechanisms of proteasome inhibitor action and resistance in cancer. Drug Resist Updat 11:164–179

    Article  CAS  PubMed  Google Scholar 

  • Mengual E, Arizti P, Rodrigo J, Giménez-Amaya JM, Castaño JG (1996) Immunohistochemical distribution and electron microscopic subcellular localization of the proteasome in the rat CNS. J Neurosci 16:6331–6341

    CAS  PubMed  Google Scholar 

  • Modem S, Badri KR, Holland TC, Reddy TR (2005) Sam68 is absolutely required for Rev function and HIV-1 production. Nucleic Acids Res 33:873–879

    Article  CAS  PubMed  Google Scholar 

  • Nandi D, Tahiliani P, Kumar A, Chandu D (2006) The ubiquitin-proteasome system. J Biosci 31:137–155

    Article  CAS  PubMed  Google Scholar 

  • Paronetto MP, Messina V, Bianchi E, Barchi M, Vogel G, Moretti C, Palombi F, Stefanini M, Geremia R, Richard S, Sette C (2009) Sam68 regulates translation of target mRNAs in male germ cells, necessary for mouse spermatogenesis. J Cell Biol 185:235–249

    Article  CAS  PubMed  Google Scholar 

  • Pena E, Berciano MT, Fernandez R, Crespo P, Lafarga M (2000) Stress-induced activation of c-Jun N-terminal kinase in sensory ganglion neurons: accumulation in nuclear domains enriched in splicing factors and distribution in perichromatin fibrils. Exp Cell Res 10:179–191

    Article  Google Scholar 

  • Pena E, Berciano MT, Fernandez R, Ojeda JL, Lafarga M (2001) Neuronal body size correlates with the number of nucleoli and Cajal bodies, and with the organization of the splicing machinery in rat trigeminal ganglion neurons. J Comp Neurol 430:250–263

    Article  CAS  PubMed  Google Scholar 

  • Rajan P, Gaughant L, Dalgliesh C, El-Sherif A, Robson CN, Leung HY, Elliott DJ (2008) Regulation of gene expression by the RNA-binding protein SAM68 in cancer. Biochem Soc Trans 36:505–507

    Article  CAS  PubMed  Google Scholar 

  • Raska I, Ochs RL, Andrade LEC, Chan EKL, Burlingame R, Peebles C, Gruol D, Tan EM (1990) Association between the nucleolus and the coiled body. J Struct Biol 104:120–127

    Article  CAS  PubMed  Google Scholar 

  • Richardson PG, Mitsiades C, Hideshima T, Anderson KC (2006a) Bortezomib: proteasome inhibition as an effective anticancer therapy. Annu Rev Med 57:33–47

    Article  CAS  PubMed  Google Scholar 

  • Richardson PG, Briemberg H, Jagannath S, Wen PY, Barlogie B, Berenson J, Singhal S, Siegel DS, Irwin D, Schuster M, Srkalovic G, Alexanian R, Rajkumar SV, Limentani S, Alsina M, Orlowski RZ, Najarian K, Esseltine D, Anderson KC, Amato AA (2006b) Frequency, characteristics, and reversibility of peripheral neuropathy during treatment of advanced multiple myeloma with bortezomib. J Clin Oncol 24:3113–3120

    Article  CAS  PubMed  Google Scholar 

  • Rouquette J, Kalland KH, Fakan S (2008) Visualization of RNA by electron microscopic in situ hybridization. In: Hancock R (ed) The nucleus: volume 2. Humana Press, Totowa, NJ, pp 403–413

    Google Scholar 

  • Shah MH, Young D, Kindler HL, Webb I, Kleiber B, Wright J, Grever M (2004) Phase II study of the proteasome inhibitor bortezomib (PS-341) in patients with metastatic neuroendocrine tumors. Clin Cancer Res 10:6111–6118

    Article  CAS  PubMed  Google Scholar 

  • Spector WS (1956) Handbook of biological data. W.B. Sanders Co., Philadelphia, p 175

    Google Scholar 

  • Szutorisz H, Georgiou A, Tora L, Dillon N (2006) The proteasome restricts permissive transcription at tissue-specific gene loci in embryonic stem cells. Cell 127:1375–1388

    Article  CAS  PubMed  Google Scholar 

  • Treier M, Staszewski LM, Bohmann D (1994) Ubiquitin-dependent c-Jun degradation in vivo is mediated by the delta domain. Cell 78:787–798

    Article  CAS  PubMed  Google Scholar 

  • Woulfe J (2008) Nuclear bodies in neurodegenerative disease. Biochim Biophys Acta 1783:2195–2206

    Article  CAS  PubMed  Google Scholar 

  • Zimmermann J, Erdmann D, Lalande I, Grossenbacher R, Noorani M, Furst P (2000) Proteasome inhibitor induced gene expression profiles reveal overexpression of transcriptional regulators ATF3, GADD153 and MAD1. Oncogene 19:2913–2920

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The authors wish to thank Raquel García-Ceballos and Saray Pereda for technical assistance. We are also grateful to Prof. E. Wahle (University of Halle, Germany) for generously providing anti-PABPN1 rabbit serum, Dr. Stéphane Richard (McGill University, Canada) for anti-Sam68 (AD1) antibody, Prof. A. I. Lamond (University of Dundee, UK) for anti-coilin antibody (204.10), and Prof. M. Carmo-Fonseca (Institute of Molecular Medicine, Lisbon) for mouse polyclonal anti-PABPN1 antibody. This work was supported by the following grants: “Dirección General de Investigación” of Spain (BFU2008-00175), and “Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED; CB06/05/0037)” from Spain.

Author information

Authors and Affiliations

  1. Department of Anatomy and Cell Biology and “Centro de Investigación Biomédica en Red sobre Enferemedades Neurodegenerativas (CIBERNED)”, Faculty of Medicine, University of Cantabria, Avd. Cardenal Herrera Oria s/n, Santander, 39011, Spain

    Iñigo Casafont, Maria T. Berciano & Miguel Lafarga

Authors
  1. Iñigo Casafont
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Maria T. Berciano
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Miguel Lafarga
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Miguel Lafarga.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Casafont, I., Berciano, M.T. & Lafarga, M. Bortezomib Induces the Formation of Nuclear poly(A) RNA Granules Enriched in Sam68 and PABPN1 in Sensory Ganglia Neurons. Neurotox Res 17, 167–178 (2010). https://doi.org/10.1007/s12640-009-9086-1

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12640-009-9086-1

Keywords

Search

Navigation