Determinants of amyloid fibril degradation by the PDZ protease HTRA1

Poepsel, Simon; Sprengel, Andreas; Saccá, Barbara; Kaschani, Farnusch; Kaiser, Markus; Gatsogiannis, Christos; Raunser, Stefan; Clausen, Tim; Ehrmann, Michael

doi:10.1038/nchembio.1931

Cited by 87 publications

(109 citation statements)

References 57 publications

(66 reference statements)

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“…5X SDS sample buffer (final concentration of 10 mM Tris, pH 6.8, 1 mM EDTA, 40 mM DTT, 0.005% Bromophenol Blue, 0.0025% Pyronin Yellow, 1% SDS, 10% sucrose) was then added to both the CSK soluble and insoluble fractions (referred to as the Triton-soluble and -insoluble fractions) and the samples were heated at 100°C for 10 minutes as well as probe sonicated for the Triton-insoluble samples. For Sarkosyl fractionation, cells were washed with PBS and harvested from 6-well plates in 400 μl lysis buffer containing 1% Sarkosyl, 10 mM Tris-HCl, 150 mM NaCl, 1 mM EGTA, and 5 mM EDTA, pH 7.4 with protease inhibitor as previously described (81). The cell suspensions were syringe-sheared with a 27G needle, followed by incubation on ice for 15 minutes, twice bath sonication for 2 minutes and incubation at RT for 20 minutes.…”

Section: Methodsmentioning

confidence: 99%

Distinct differences in prion-like seeding and aggregation between Tau protein variants provide mechanistic insights into tauopathies

Strang¹,

Croft²,

Sorrentino³

et al. 2018

Journal of Biological Chemistry

120

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The accumulation of aberrantly aggregated microtubule-associated protein tau (MAPT) 1 defines a spectrum of tauopathies, including Alzheimer's disease. Mutations in the MAPT gene cause frontotemporal dementia with Parkinsonism linked to chromosome 17 (FTDP-17), characterized by neuronal pathological tau inclusions in the form of neurofibrillary tangles and Pick bodies, and in some cases glial tau pathology. Increasing evidence points to the importance of prion-like seeding as a mechanism for the pathological spread in tauopathy and other neurodegenerative diseases. Herein, using a cell culture model, we examined a multitude of genetic FTDP-17 tau variants for their ability to be seeded by exogenous tau fibrils. Our findings revealed stark differences between FTDP-17 tau variants in their ability to be seeded, with variants at proline 301 and serine 320 showing robust aggregation with seeding. Similarly, we elucidated the importance of certain tau protein regions and unique residues, including the role of proline 301 in inhibiting tau aggregation. We also revealed potential barriers in cross-seeding between 3-repeat and 4-repeat tau isoforms. Overall, these differences alluded to potential mechanistic differences between wild-type and FTDP-17 tau variants, as well as different tau isoforms, in influencing tau aggregation. Furthermore, by combining two FTDP-17 tau variants (either P301L or P301S with S320F), we generated aggressive models of tauopathy that does not require exogenous seeding. These models will allow for rapid screening of potential therapeutics to alleviate tau aggregation without the need for exogenous tau fibrils. Together, these studies provide novel insights in the molecular determinants that modulate tau aggregation.Tauopathies are a spectrum of neurodegenerative diseases characterized by the presence of pathological inclusions composed of aberrantly aggregated and hyperphosphorylated microtubule-associated protein tau (MAPT). Tauopathies are pathologically and phenotypically diverse, and include Alzheimer's disease (AD), progressive supranuclear palsy, corticobasal degeneration, chronic traumatic encephalopathy, Pick's disease, and frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17)(1-4). Tau is abundant in neurons and expressed at lower levels in glia (1, 6, 7), and primarily stabilizes microtubules (MTs) among other diverse physiological functions (1,5,8,9). Six different isoforms of tau, ranging from 352 to 441 amino acids, are expressed in the adult human brain as a result of alternative splicing (10, 11). Differential splicing results in the inclusion or exclusion of the R2 MT-binding repeat, producing tau isoforms with either three (3R) or four (4R) repeats, respectively, and one or two N-terminal inserts (Figure 1).Over 50 mutations have been identified in the MAPT gene in families with 2,4,12 Tau Mutants and Seeding 2 disease, and patients typically experience disease onset at ~49 years of age with an average disease duration of 8.5 years (13). Certai...

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Section: Methodsmentioning

confidence: 99%

Distinct differences in prion-like seeding and aggregation between Tau protein variants provide mechanistic insights into tauopathies

Strang¹,

Croft²,

Sorrentino³

et al. 2018

Journal of Biological Chemistry

120

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“…Furthermore, HtrA1 colocalizes with amyloid deposits in human brain samples [36]. HtrA1 also degrades aggregated and fibrillar tau protein; neuronal cells and patient brains accumulate less tau, neurofibrillary tangles, and neuritic plaques, respectively, when HtrA1 is expressed at elevated levels [43,44]. These data suggest that HtrA1 performs regulated proteolysis during protein quality control which protects the brain against accumulation of amyloid deposits and tau neurofibrillary tangles e the hallmarks of the Alzheimer's disorder.…”

Section: Human Htra Proteins In Cellular Physiology and Pathogenesismentioning

confidence: 80%

Structural insights into the activation mechanisms of human HtrA serine proteases

Żurawa-Janicka

Wenta

Jarzab

et al. 2017

Archives of Biochemistry and Biophysics

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“…In vitro biochemical studies into the regulation of Tau led to the identification of HtrA1 as a Tau degrading enzyme. HtrA1 has recently been reported to have a non-proteolytic activity that enables it to disentangle Tau neurofibrillary tangles, suggesting that it can regulate the levels and aggregation of Tau 34 .…”

Section: Resultsmentioning

confidence: 99%

HtrA1 Proteolysis of ApoE In Vitro Is Allele Selective

et al. 2016

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Apolipoprotein E (ApoE) belongs to a large class of proteins that solubilize lipids for physiological transport. Humans have three different APOE alleles—APOE ε2, APOE ε3 and APOE ε4—and genetic studies identified ApoE4 as the strongest genetic risk factor for Alzheimer’s disease (AD). People who are homozygous for ApoE4 (i.e. ApoE4/E4) are an order of magnitude more likely to develop late-onset AD (LOAD) than ApoE3/E3 carriers. Several differences between ApoE3 and ApoE4 may contribute to AD including the observation that ApoE4 is degraded to a greater extent than ApoE3 in the human brain. Experiments with high-temperature requirement serine peptidase A1 (HtrA1), which is found in the nervous system, demonstrate that HtrA1 is an allele-selective ApoE-degrading enzyme that degrades ApoE4 more quickly than ApoE3. This activity is specific to HtrA1, as similar assays with HtrA2 showed minimal ApoE4 proteolysis and trypsin had no preference between ApoE4 and ApoE3. HtrA1 has also been reported to cleave the tau protein (Tau) and the amyloid protein precursor (APP) to hinder the formation of toxic amyloid deposits associated with AD. Competition assays with ApoE4, ApoE3, and Tau revealed that ApoE4 inhibits Tau degradation. Thus, the identification of ApoE4 as an in vitro HtrA1 substrate suggests a potential biochemical mechanism that links ApoE4 regulation of AD proteins such as Tau.

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Determinants of amyloid fibril degradation by the PDZ protease HTRA1

Cited by 87 publications

References 57 publications

Distinct differences in prion-like seeding and aggregation between Tau protein variants provide mechanistic insights into tauopathies

Distinct differences in prion-like seeding and aggregation between Tau protein variants provide mechanistic insights into tauopathies

Structural insights into the activation mechanisms of human HtrA serine proteases

HtrA1 Proteolysis of ApoE In Vitro Is Allele Selective

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