Oligomeric and polymeric aggregates formed by proteins containing expanded polyglutamine

Iuchi, Shiro; Hoffner, Guylaine; Verbeke, Philippe; Djian, Philippe; Green, H.

doi:10.1073/pnas.0437660100

Cited by 63 publications

(56 citation statements)

References 51 publications

(42 reference statements)

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“…To date, there has been no conclusive evidence for the existence of soluble polyglutamine protein oligomers in patients or animal models, nor any evidence that these species actually underlie toxicity in vivo. Green and colleagues (33) reported large formic acid non-dissociable "oligomers" in Huntington disease brain but did not clearly determine the nature of their protein constituents or physical properties. Establishing that oligomers are a key pathologic species in vivo would advance our basic understanding of disease mechanisms and help in developing new therapies and in monitoring disease onset and progression.…”

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confidence: 99%

Soluble Androgen Receptor Oligomers Underlie Pathology in a Mouse Model of Spinobulbar Muscular Atrophy

Chevalier-Larsen

Merry

et al. 2007

Journal of Biological Chemistry

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In polyglutamine diseases such as X-linked spinobulbar muscular atrophy (SBMA), it is unknown whether the toxic form of the protein is an insoluble or soluble aggregate or a monomer. We have addressed this question by studying a full-length androgen receptor (AR) mouse model of SBMA. We used biochemistry and atomic force microscopy to immunopurify oligomers soluble after ultracentrifugation that are comprised of a single ϳ50-kDa N-terminal polyglutamine-containing AR fragment. AR oligomers appeared several weeks prior to symptom onset, were distinct and temporally dissociated from intranuclear inclusions, and disappeared rapidly after castration, which halts disease. This is the first demonstration of soluble AR oligomers in vivo and suggests that they underlie neurodegeneration in SBMA.Polyglutamine diseases such as X-linked spinobulbar muscular atrophy (SBMA) 2 derive from CAG codon repeats that exceed a crucial length. This creates elongated polyglutamine tracts in affected proteins and progressive neurologic dysfunction and neurodegeneration (1). SBMA results from expanded glutamine repeats in the N terminus of the androgen receptor (AR) protein (2, 3). It is characterized by slowly progressive lower motor neuron degeneration and mild sensory neuronopathy that predominates in males (4, 5) due to exposure to androgens (6, 7). Large intracellular inclusions containing AR are observed throughout the body and may be localized in the cytoplasm or nucleus, whereas large intranuclear inclusions accumulate preferentially in motor neurons (8 -10). Pathologically expanded polyglutamine protein fragments are clearly prone to misfolding and aggregation in vitro that respects the tight length-dependent disease threshold seen in vivo (11,12). Thus, protein misfolding and/or aggregation are very likely to underlie pathogenesis of SBMA and other polyglutamine diseases. In many experimental systems, however, large intracellular inclusions can be dissociated from neuronal toxicity, indicating that these structures are probably not the primary pathologic species (13)(14)(15)(16)(17)(18)(19).If large inclusions are not pathogenic, what then is the toxic species? Most cellular and animal studies indicate that expanded proteins either exist as monomers or accumulate as large macromolecular complexes that are unable to enter an SDS-polyacrylamide gel (20 -23). However, in vitro studies indicate clearly that polyglutamine proteins, like other amyloidogenic proteins, can form small, ordered oligomers (11, 24 -27), and the existence of toxic, submicroscopic aggregates or oligomers has been invoked as a cause of polyglutamine toxicity in vivo (27,28). The operative definition of such species has varied. We define oligomers here as submacromolecular structures (i.e. soluble after high speed centrifugation) comprised of ordered polyglutamine aggregates. As previously proposed by Taylor et al. (28), we contrast protein oligomers, which are defined primarily via biochemistry, from intracellular inclusions, which are defined via histopath...

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confidence: 99%

Soluble Androgen Receptor Oligomers Underlie Pathology in a Mouse Model of Spinobulbar Muscular Atrophy

Chevalier-Larsen

Merry

et al. 2007

Journal of Biological Chemistry

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“…An important characteristic of the protein aggregates found in Huntington's, Parkinson's, and Alzheimer's disease is their insolubility in high concentrations (≥2%) of SDS. 17,19,51 We found that the GFP N104 proteins were trapped at the interface of the stacking and separating gels in SDS/PAGE, even after the cell lysates were boiled in a buffer containing 2% SDS (data not shown), suggesting the presence of SDS-insoluble aggregates. To further determine if GFP N104 could form SDS-resistant aggregates, we used a cellulose acetate membrane based filter trap assay to detect SDS-insoluble protein oligomers that were larger than 0.2 mm in solution.…”

Section: Resultsmentioning

confidence: 95%

“…14 Consistent with this notion, insertion of a pathogenic length glutamine tract in the hypoxanthine phosphoribosyltransferase (Hprt) protein in a knock-in mouse model results in protein aggregation, behavioral abnormalities, and shortened life spans similar to the phenotypes observed in the mouse model of Huntington's Disease. 15,16 Polyglutamine proteins have been shown to exist in different oligomeric states both in vitro and in vivo, [17][18][19] and the roles of these different species of aggregates in pathogenesis are being actively investigated.…”

Section: Introductionmentioning

confidence: 99%

Protein Aggregation and Polyasparagine-Mediated Cellular Toxicity inSaccharomyces cerevisiae

Peters

Huang

2007

Prion

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It is well established that protein aggregation is associated with many neurodegenerative disorders including polyglutamine diseases, but a mechanistic understanding of the role of protein aggregates in the disease pathogenesis remains elusive. Previously thought to be the cause of cellular toxicity such as cellular dysfunction and cell death, protein aggregation is now proposed to serve a protective role by sequestering toxic oligomers from interfering with essential physiological processes. To investigate the relationship between protein aggregation and cellular toxicity, we have characterized and compared the effects of two GFP-fusion proteins that form aggregates in Saccharomyces cerevisiae, one with a polyasparagine repeat (GFP N104 ) and one without (GFP C ). Although both proteins can form microscopically visible GFP-positive aggregates, only the GFP N104 -containing aggregates exhibit morphological and biochemical characteristics that resemble the aggregates formed by mutant huntingtin in yeast cells. Formation of both the GFP C and GFP N104 aggregates depends on microtubules, while only the GFP N104 aggregate requires the chaperone Hsp104 and the prion Rnq1 and is resistant to SDS. Although no microscopically visible GFP N104 aggregates were observed in the hsp104D and rnq1D mutant cells, SDS-insoluble aggregates can still be detected by the filter trap assay. These observations argue that the GFP N104 -containing aggregates can exist in at least two distinct states in vivo. We also show that a nucleus-targeted GFP N104 interferes with transcription from two SAGA-dependant promoters and results in a decrease in cell viability. Overall, the results imply that the GFP N104 protein behaves similarly to the mutant huntingtin in yeast cells and provides a new model for investigating the interplay between protein aggregates and the associated phenotypes.

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“…However, the given reasoning for the pathological sequence of events observed because of aggregation of these mutated proteins is still debated. Two widely believed mechanisms of aggregate formation are either via β-pleated sheets [4,5] or by transglutaminase (TG) dependent covalent incorporation of polyQ proteins into the aggregates [6,7].…”

Section: Introductionmentioning

confidence: 99%

Role of tissue transglutaminase type 2 in calbindin–D28k interaction with ataxin-1

Vig

Wei

Shao

et al. 2007

Neuroscience Letters

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Spinocerebellar ataxia-1 (SCA1) is caused by the expansion of a polyglutamine repeats within the disease protein, ataxin-1. The mutant ataxin-1 precipitates as large intranuclear aggregates in the affected neurons. These aggregates may protect neurons from mutant protein and/or trigger neuronal degeneration by encouraging recruitment of other essential proteins. Our previous studies have shown that calcium binding protein calbindin-D28k (CaB) associated with SCA1 pathogenesis is recruited to ataxin-1 aggregates in Purkinje cells of SCA1 mice. Since our recent findings suggest that tissue transglutaminase 2 (TG2) may be involved in cross-linking and aggregation of ataxin-1, the present study was initiated to determine if TG2 has any role in CaB-ataxin-1 interaction. The guinea pig TG2 covalently cross-linked purified rat brain CaB. Time dependent progressive increase in aggregation produced large multimers, which stayed on top of the gel. CaB interaction with ataxin-1 was studied using HeLa cell lysates expressing GFP and GFP tagged ataxin-1 with normal and expanded polyglutamine repeats (Q2, Q30 and Q82). The reaction products were analyzed by Western blots using anti-polyglutamine, CaB or GFP antibodies. CaB interacted with ataxin-1 independent of TG2 as the protein-protein cross-linker DSS stabilized CaB-ataxin-1 complex. TG2 cross-linked CaB preferentially with Q82 ataxin-1. The cross-linking was inhibited with EGTA or TG2 inhibitor cystamine. The present data indicate that CaB may be a TG2 substrate. In addition, aggregates of mutant ataxin-1 may recruit CaB via TG2 mediated covalent cross-linking, further supporting the argument that ataxin-1 aggregates may be toxic to neurons.

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Oligomeric and polymeric aggregates formed by proteins containing expanded polyglutamine

Cited by 63 publications

References 51 publications

Soluble Androgen Receptor Oligomers Underlie Pathology in a Mouse Model of Spinobulbar Muscular Atrophy

Soluble Androgen Receptor Oligomers Underlie Pathology in a Mouse Model of Spinobulbar Muscular Atrophy

Protein Aggregation and Polyasparagine-Mediated Cellular Toxicity inSaccharomyces cerevisiae

Role of tissue transglutaminase type 2 in calbindin–D28k interaction with ataxin-1

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