Oxidative stress promotes mutant huntingtin aggregation and mutant huntingtin-dependent cell death by mimicking proteasomal malfunction

https://doi.org/10.1016/j.bbrc.2006.01.136Get rights and content

Abstract

Huntington’s disease (HD) is a familial neurodegenerative disorder caused by an abnormal expansion of CAG repeats in the coding region of huntingtin gene. A major hallmark of HD is the proteolytic production of N-terminal fragments of huntingtin containing polyglutamine repeats that form ubiquitinated aggregates in the nucleus and cytoplasm of the affected neurons. However, the mechanism by which the mutant huntingtin causes neurodegeneration is not well understood. Here, we found that oxidative stimuli enhance the polyglutamine-expanded truncated N-terminal huntingtin (mutant huntingtin) aggregation and mutant huntingtin-induced cell death. Oxidative stimuli also lead to rapid proteasomal dysfunction in the mutant huntingtin expressing cells as compared to normal glutamine repeat expressing cells. Overexpression of Cu/Zn superoxide dismutase (SOD1), Hsp40 or Hsp70 reverses the oxidative stress-induced proteasomal malfunction, mutant huntingtin aggregation, and death of the mutant huntingtin expressing cells. Finally, we show the higher levels of expression of SOD1 and DJ-1 in the mutant huntingtin expressing cells. Our result suggests that oxidative stress-induced proteasomal malfunction might be linked with mutant huntingtin-induced cell death.

Section snippets

Materials and methods

Materials. Proteasome substrate, MG132, lactacystin, N-acetyl cysteine, rabbit polyclonal ubiquitin antibody, mouse monoclonal β-tubulin, and all cell culture reagents were obtained from Sigma. LipofectAMINE 2000 and ponasterone A were obtained from Invitrogen, mouse monoclonal anti-HA was obtained from Roche, rabbit polyclonal anti-SOD1 and DJ-1 and mouse monoclonal anti-Hsp70 were from Santa Cruz, and AP-conjugated anti-mouse and anti-rabbit IgG were from Vector laboratories. Ubiquitin

Oxidative stimuli enhances mutant huntingtin-induced cell death and mutant huntingtin aggregation

To test the possible effect of oxidative stress on the expanded polyglutamine protein-induced cell death we used stable and inducible cell lines that express tNhtt fused with EGFP containing 16 and 150 polyglutamine residues. The cell lines were named HD 16Q and HD 150Q and their corresponding expressed proteins were named tNhtt-16Q and tNhtt-150Q. The cell lines were induced with 1 μM ponasterone A for 2 days and then exposed to different doses of H2O2 for 5 h. The cell lines were induced for 2

Discussion

Ample evidence indicates that oxidative stress and energy defect may play an important role in the pathogenesis of polyglutamine diseases including HD [9], [10], [11], [12], [13]. Recent studies have also demonstrated the involvement of proteasome impairment in vast majority of neurodegenerative disorders including polyglutamine diseases [7], [8]. But the relationship between oxidative stress and proteasomal malfunction in the polyglutamine diseases is not known. In the present investigation,

Acknowledgments

This work was supported by the Department of Biotechnology, Government of India. P.D. and A.M. were supported by research fellowship from Council of Scientific and Industrial Research, Government of India.

References (27)

  • N.F. Bence et al.

    Impairment of the ubiquitin–proteasome system by protein aggregation

    Science

    (2001)
  • S.E. Browne et al.

    Oxidative damage and metabolic dysfunction in Huntington’s disease: selective vulnerability of the basal ganglia

    Ann. Neurol.

    (1997)
  • S.E. Browne et al.

    Oxidative stress in Huntington’s disease

    Brain Pathol.

    (1999)
  • Cited by (107)

    View all citing articles on Scopus
    View full text