Architecture of Polyglutamine-containing Fibrils from Time-resolved Fluorescence Decay

Röthlein, Christoph; Miettinen, Markus S.; Borwankar, Tejas; Bürger, Jörg; Mielke, Thorsten; Kumke, Michael U.; Ignatova, Zoya

doi:10.1074/jbc.m114.581991

Cited by 9 publications

(6 citation statements)

References 37 publications

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“…The oligomerization and toxicity of expanded Htt-polyQ is inhibited by molecular chaperones, such as Hsp70–Hsp40, CCT/TRiC, and Prefoldin [ 22 , 23 , 25 , 26 , 27 , 28 ]. Typically, intermolecular FRET detection of polyQ-protein oligomerization is based on stoichiometric labeling of polyQ-protein with a donor- and acceptor-fluorescent tag and is determined by fluorescence intensity ratio between the donor and acceptor [ 23 , 24 , 29 ]. After establishing the increase in the FRET efficiency due to oligomerization of expanded Htt-polyQ following incubation and/or agitation of the labeled proteins in vitro , one can determine the rate of decrease in the FRET efficiency following the addition of purified molecular chaperones [ 23 ].…”

Section: Spectroscopic Imaging For Elucidation Of the Mechanism Ofmentioning

confidence: 99%

Conformational Analysis of Misfolded Protein Aggregation by FRET and Live-Cell Imaging Techniques

Kitamura

Nagata

2015

IJMS

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Cellular homeostasis is maintained by several types of protein machinery, including molecular chaperones and proteolysis systems. Dysregulation of the proteome disrupts homeostasis in cells, tissues, and the organism as a whole, and has been hypothesized to cause neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS) and Huntington’s disease (HD). A hallmark of neurodegenerative disorders is formation of ubiquitin-positive inclusion bodies in neurons, suggesting that the aggregation process of misfolded proteins changes during disease progression. Hence, high-throughput determination of soluble oligomers during the aggregation process, as well as the conformation of sequestered proteins in inclusion bodies, is essential for elucidation of physiological regulation mechanism and drug discovery in this field. To elucidate the interaction, accumulation, and conformation of aggregation-prone proteins, in situ spectroscopic imaging techniques, such as Förster/fluorescence resonance energy transfer (FRET), fluorescence correlation spectroscopy (FCS), and bimolecular fluorescence complementation (BiFC) have been employed. Here, we summarize recent reports in which these techniques were applied to the analysis of aggregation-prone proteins (in particular their dimerization, interactions, and conformational changes), and describe several fluorescent indicators used for real-time observation of physiological states related to proteostasis.

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Section: Spectroscopic Imaging For Elucidation Of the Mechanism Ofmentioning

confidence: 99%

Conformational Analysis of Misfolded Protein Aggregation by FRET and Live-Cell Imaging Techniques

Kitamura

Nagata

2015

IJMS

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“…They could pinpoint the dynamic structure of this oligomer to a patchwork of amino acid segments which fold locally before the transition into highly ordered amyloid filaments. Likewise, Röthlein et al employed time resolved fluorescence and computational calculations to obtain structural views of an extremely unstable huntingtin (htt) amyloid filament (Röthlein et al, 2014 ).…”

Section: Amyloid Aggregationmentioning

confidence: 99%

Legal but lethal: functional protein aggregation at the verge of toxicity

Falsone

2015

Front. Cell. Neurosci.

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Many neurodegenerative disorders are linked to irreversible protein aggregation, a process that usually comes along with toxicity and serious cellular damage. However, it is emerging that protein aggregation can also serve for physiological purposes, as impressively shown for prions. While the aggregation of this protein family was initially considered exclusively toxic in mammalians organisms, it is now almost clear that many other proteins adopt prion-like attributes to rationally polymerize into higher order complexes with organized physiologic roles. This implies that cells can tolerate at least in some measure the accumulation of inherently dangerous protein aggregates for functional profit. This review summarizes currently known strategies that living organisms adopt to preserve beneficial aggregation, and to prevent the catastrophic accumulation of toxic aggregates that frequently accompany neurodegeneration.

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“…For all of these polyQ diseases, it is the aggregate formation that leads to the disease. Longer polyQ segments are conducive to β-sheet formation 15 16 17 that are recognized by chaperone proteins as misfolded and produce the aggregates characteristic of the disease 5 18 19 . Other aggregate forming proteins can also become incorporated into Htt-polyQ aggregates 20 21 .…”

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confidence: 99%

Multiple discrete soluble aggregates influence polyglutamine toxicity in a Huntington’s disease model system

Wang

Laue

et al. 2016

Sci Rep

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Huntington’s disease (HD) results from expansions of polyglutamine stretches (polyQ) in the huntingtin protein (Htt) that promote protein aggregation, neurodegeneration, and death. Since the diversity and sizes of the soluble Htt-polyQ aggregates that have been linked to cytotoxicity are unknown, we investigated soluble Htt-polyQ aggregates using analytical ultracentrifugation. Soon after induction in a yeast HD model system, non-toxic Htt-25Q and cytotoxic Htt-103Q both formed soluble aggregates 29S to 200S in size. Because current models indicate that Htt-25Q does not form soluble aggregates, reevaluation of previous studies may be necessary. Only Htt-103Q aggregation behavior changed, however, with time. At 6 hr mid-sized aggregates (33S to 84S) and large aggregates (greater than 100S) became present while at 24 hr primarily only mid-sized aggregates (20S to 80S) existed. Multiple factors that decreased cytotoxicity of Htt-103Q (changing the length of or sequences adjacent to the polyQ, altering ploidy or chaperone dosage, or deleting anti-aging factors) altered the Htt-103Q aggregation pattern in which the suite of mid-sized aggregates at 6 hr were most correlative with cytotoxicity. Hence, the amelioration of HD and other neurodegenerative diseases may require increased attention to and discrimination of the dynamic alterations in soluble aggregation processes.

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Architecture of Polyglutamine-containing Fibrils from Time-resolved Fluorescence Decay

Cited by 9 publications

References 37 publications

Conformational Analysis of Misfolded Protein Aggregation by FRET and Live-Cell Imaging Techniques

Conformational Analysis of Misfolded Protein Aggregation by FRET and Live-Cell Imaging Techniques

Legal but lethal: functional protein aggregation at the verge of toxicity

Multiple discrete soluble aggregates influence polyglutamine toxicity in a Huntington’s disease model system

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