In Vitro and in Vivo Aggregation of a Fragment of Huntingtin Protein Directly Causes Free Radical Production

• Published in: The Journal of biological chemistry Vol. 286; no. 52; pp. 44512 - 44520
• Main Authors: Hands, Sarah, Sajjad, Mohammad U., Newton, Michael J., Wyttenbach, Andreas
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 30-12-2011; American Society for Biochemistry and Molecular Biology
• Subjects: Cell Death; Cell Line; Humans; Huntingtin Protein; Huntington Disease - genetics; Huntington Disease - metabolism; Hydrogen Peroxide - metabolism; Microscopy, Atomic Force; Models, Biological; Nerve Tissue Proteins - chemistry; Nerve Tissue Proteins - genetics; Nerve Tissue Proteins - metabolism; Neurobiology; Neurodegeneration; Neurodegenerative Diseases; Nuclear Proteins - chemistry; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Peptides - chemistry; Peptides - genetics; Peptides - metabolism; Polyglutamine; Polyglutamine Disease; Protein Aggregation; Protein Misfolding; Protein Multimerization; Protein Structure, Quaternary; Single-Chain Antibodies - chemistry; Neurodegenerative Diseases; Protein Misfolding; Polyglutamine Disease; Polyglutamine; Protein Aggregation; Neurodegeneration
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.307587

Neurodegenerative diseases are characterized by intra- and/or extracellular protein aggregation and oxidative stress. Intense attention has been paid to whether protein aggregation itself contributes to abnormal production of free radicals and ensuing cellular oxidative damage. Although this question has been investigated in the context of extracellular protein aggregation, it remains unclear whether protein aggregation inside cells alters the redox homeostasis. To address this, we have used in vitro and in vivo (cellular) models of Huntington disease, one of nine polyglutamine (poly(Q)) disorders, and examined the causal relationship among intracellular protein aggregation, reactive oxygen species (ROS) production, and toxicity. Live imaging of cells expressing a fragment of huntingtin (httExon1) with a poly(Q) expansion shows increased ROS production preceding cell death. ROS production is poly(Q) length-dependent and not due to the httExon 1 flanking sequence. Aggregation inhibition by the MW7 intrabody and Pgl-135 treatment abolishes ROS production, showing that increased ROS is caused by poly(Q) aggregation itself. To examine this hypothesis further, we determined whether aggregation of poly(Q) peptides in vitro generated free radicals. Monitoring poly(Q) protein aggregation using atomic force microscopy and hydrogen peroxide (H2O2) production over time in parallel we show that oligomerization of httEx1Q53 results in early generation of H2O2. Inhibition of poly(Q) oligomerization by the single chain antibody MW7 abrogates H2O2 formation. These results demonstrate that intracellular protein aggregation directly causes free radical production, and targeting potentially toxic poly(Q) oligomers may constitute a therapeutic target to counteract oxidative stress in poly(Q) diseases.
Neurodegenerative diseases are associated with intracellular protein aggregation and free radical damage.
Protein aggregation of polyglutamine-containing proteins directly causes free radical production in vitro and within cells.
Protein aggregation during polyglutamine diseases could be targeted to prevent oxidative stress.
Intracellular protein aggregation during chronic neurodegeneration is closely linked to abnormal production of free radicals.