Structural Mechanisms of Mutant Huntingtin Aggregation Suppression by the Synthetic Chaperonin-like CCT5 Complex Explained by Cryoelectron Tomography

Darrow, Michele C.; Sergeeva, Oksana A.; Isas, J. Mario; Galaz-Montoya, Jesús G.; King, Jonathan; Langen, Ralf; Schmid, Michael F.; Chiu, Wah

doi:10.1074/jbc.m115.655373

Cited by 40 publications

(43 citation statements)

References 47 publications

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“…mHtt-Ex1 fibrils have a characteristic architecture, in which frayed fibril ends branch out from a bundled central core (Figure 2A, Figure 2—figure supplement 1A) (Bugg et al, 2012; Darrow et al, 2015; Shahmoradian et al, 2013). For the ∆N mHtt variant, we observed dramatically fewer fibrils, consistent with its lower amyloid aggregation propensity (Figure 1).…”

Section: Resultsmentioning

confidence: 99%

Control of the structural landscape and neuronal proteotoxicity of mutant Huntingtin by domains flanking the polyQ tract

Shen

Calamini

Fauerbach

et al. 2016

eLife

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108

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Many neurodegenerative diseases are linked to amyloid aggregation. In Huntington’s disease (HD), neurotoxicity correlates with an increased aggregation propensity of a polyglutamine (polyQ) expansion in exon 1 of mutant huntingtin protein (mHtt). Here we establish how the domains flanking the polyQ tract shape the mHtt conformational landscape in vitro and in neurons. In vitro, the flanking domains have opposing effects on the conformation and stabilities of oligomers and amyloid fibrils. The N-terminal N17 promotes amyloid fibril formation, while the C-terminal Proline Rich Domain destabilizes fibrils and enhances oligomer formation. However, in neurons both domains act synergistically to engage protective chaperone and degradation pathways promoting mHtt proteostasis. Surprisingly, when proteotoxicity was assessed in rat corticostriatal brain slices, either flanking region alone sufficed to generate a neurotoxic conformation, while the polyQ tract alone exhibited minimal toxicity. Linking mHtt structural properties to its neuronal proteostasis should inform new strategies for neuroprotection in polyQ-expansion diseases.DOI: http://dx.doi.org/10.7554/eLife.18065.001

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Section: Resultsmentioning

confidence: 99%

Control of the structural landscape and neuronal proteotoxicity of mutant Huntingtin by domains flanking the polyQ tract

Shen

Calamini

Fauerbach

et al. 2016

eLife

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108

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“…CryoET can also be used to structurally characterize pleomorphic (Harris et al 2006) and disordered specimens, such as amyloid aggregates (Darrow et al 2015; Shahmoradian & Galaz-Montoya et al 2013), as well as complexes exhibiting conformational and compositional heterogeneity (Dai et al 2013). Due to mechanical limitations of the electron microscope and the slab geometry of frozen-hydrated specimens, imaging is typically limited to a maximum useful tilt of ±60°, leaving a region of angular space from which micrographs of the specimen cannot be recorded.…”

Section: Introductionmentioning

confidence: 99%

Alignment algorithms and per-particle CTF correction for single particle cryo-electron tomography

Galaz-Montoya

Hecksel²,

Baldwin

et al. 2016

Journal of Structural Biology

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Single particle cryo-electron tomography (cryoSPT) extracts features from cryo-electron tomograms, followed by 3D classification, alignment and averaging to generate improved 3D density maps of such features. Robust methods to correct for the contrast transfer function (CTF) of the electron microscope are necessary for cryoSPT to reach its resolution potential. Many factors can make CTF correction for cryoSPT challenging, such as lack of eucentricity of the specimen stage, inherent low dose per image, specimen charging, beam-induced specimen motions, and defocus gradients resulting both from specimen tilting and from unpredictable ice thickness variations. Current CTF correction methods for cryoET make at least one of the following assumptions: that the defocus at the center of the image is the same across the images of a tiltseries, that the particles all lie at the same Z-height in the embedding ice, and/or that the specimen grid and carbon support are flat. These experimental conditions are not always met. We have developed a CTF correction algorithm for cryoSPT without making any of the aforementioned assumptions. We also introduce speed and accuracy improvements and a higher degree of automation to the subtomogram averaging algorithms available in EMAN2. Using motion-corrected images of isolated virus particles as a benchmark specimen, recorded with a DE20 direct detection camera, we show that our CTF correction and subtomogram alignment routines can yield subtomogram averages close to 4/5 Nyquist frequency of the detector under our experimental conditions.

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“…More remarkable, following exogenous delivery, the substrate-binding apical domain of CCT1, ApiCCT1, entered the cytosol and nucleus of cells and suppressed mHTT aggregation (26). In addition, cryoelectron tomography uncovered that another individual subunit, CCT5, caps fibrils and encapsulates oligomers to inhibit mHTT aggregation (27).…”

Section: Significancementioning

confidence: 99%

TRiC subunits enhance BDNF axonal transport and rescue striatal atrophy in Huntington’s disease

Zhao

Chen

Han

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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101

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Corticostriatal atrophy is a cardinal manifestation of Huntington's disease (HD). However, the mechanism(s) by which mutant huntingtin (mHTT) protein contributes to the degeneration of the corticostriatal circuit is not well understood. We recreated the corticostriatal circuit in microfluidic chambers, pairing cortical and striatal neurons from the BACHD model of HD and its WT control. There were reduced synaptic connectivity and atrophy of striatal neurons in cultures in which BACHD cortical and striatal neurons were paired. However, these changes were prevented if WT cortical neurons were paired with BACHD striatal neurons; synthesis and release of brain-derived neurotrophic factor (BDNF) from WT cortical axons were responsible. Consistent with these findings, there was a marked reduction in anterograde transport of BDNF in BACHD cortical neurons. Subunits of the cytosolic chaperonin T-complex 1 (TCP-1) ring complex (TRiC or CCT for chaperonin containing TCP-1) have been shown to reduce mHTT levels. Both CCT3 and the apical domain of CCT1 (ApiCCT1) decreased the level of mHTT in BACHD cortical neurons. In cortical axons, they normalized anterograde BDNF transport, restored retrograde BDNF transport, and normalized lysosomal transport. Importantly, treating BACHD cortical neurons with ApiCCT1 prevented BACHD striatal neuronal atrophy by enhancing release of BDNF that subsequently acts through tyrosine receptor kinase B (TrkB) receptor on striatal neurons. Our findings are evidence that TRiC reagentmediated reductions in mHTT enhanced BDNF delivery to restore the trophic status of BACHD striatal neurons., a genetically defined progressive neurodegenerative disorder, is characterized by abnormal involuntary movements, cognitive disability, and psychiatric symptoms (1). The pathogenesis of HD is due to an expanded CAG repeat in the huntingtin (HTT) gene, which encodes the protein Htt. Mutant Htt (mHTT) features an increased number of glutamines (Q > 36) near the N terminus. The striking atrophy and loss of striatal medium-sized spiny neurons (MSNs) in the HD brain are accompanied by atrophy of the cortex (2). The mechanisms by which mHTT induces dysfunction and death of neurons are actively explored (3). It has been shown that mHTT has an impact on axonal trafficking and signaling, synaptic function, gene expression, mitochondrial function, calcium homeostasis, and proteostasis (4, 5). Thus, mHTT plays a central role in HD pathogenesis, an assertion fully supported by recent studies in the BACHD model of HD (6).One productive line of inquiry regarding HD pathogenesis focuses on brain-derived neurotrophic factor (BDNF), which sustains neurons of the corticostriatal circuit (7-10). In the corticostriatal circuit, BDNF is produced in cortical neurons and is transported anterogradely in axons before secretion in striatum. Released BDNF binds to tyrosine receptor kinase B (TrkB) on striatal MSNs, resulting in BDNF-mediated signaling. Because MSNs do not produce BDNF, they are largely dependent on cortical neurons for i...

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Structural Mechanisms of Mutant Huntingtin Aggregation Suppression by the Synthetic Chaperonin-like CCT5 Complex Explained by Cryoelectron Tomography

Cited by 40 publications

References 47 publications

Control of the structural landscape and neuronal proteotoxicity of mutant Huntingtin by domains flanking the polyQ tract

Control of the structural landscape and neuronal proteotoxicity of mutant Huntingtin by domains flanking the polyQ tract

Alignment algorithms and per-particle CTF correction for single particle cryo-electron tomography

TRiC subunits enhance BDNF axonal transport and rescue striatal atrophy in Huntington’s disease

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