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Rescue from tau-induced neuronal dysfunction produces insoluble tau oligomers.

Cowan CM, Quraishe S, Hands S, Sealey M, Mahajan S, Allan DW, Mudher A - Sci Rep (2015)

Bottom Line: Moreover, contrary to common belief, these tau oligomers were neither highly phosphorylated, and nor did they contain beta-pleated sheet structure.Whether these are inert or actively protective remains to be established.Nevertheless, this has wide implications for emerging therapeutic strategies such as those that target dissolution of tau oligomers as they may be ineffective or even counterproductive unless they act on the relevant toxic oligomeric tau species.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biological Sciences, University of Southampton, Southampton, SO17 1BJ, UK.

ABSTRACT
Aggregation of highly phosphorylated tau is a hallmark of Alzheimer's disease and other tauopathies. Nevertheless, animal models demonstrate that tau-mediated dysfunction/toxicity may not require large tau aggregates but instead may be caused by soluble hyper-phosphorylated tau or by small tau oligomers. Challenging this widely held view, we use multiple techniques to show that insoluble tau oligomers form in conditions where tau-mediated dysfunction is rescued in vivo. This shows that tau oligomers are not necessarily always toxic. Furthermore, their formation correlates with increased tau levels, caused intriguingly, by either pharmacological or genetic inhibition of tau kinase glycogen-synthase-kinase-3beta (GSK-3β). Moreover, contrary to common belief, these tau oligomers were neither highly phosphorylated, and nor did they contain beta-pleated sheet structure. This may explain their lack of toxicity. Our study makes the novel observation that tau also forms non-toxic insoluble oligomers in vivo in addition to toxic oligomers, which have been reported by others. Whether these are inert or actively protective remains to be established. Nevertheless, this has wide implications for emerging therapeutic strategies such as those that target dissolution of tau oligomers as they may be ineffective or even counterproductive unless they act on the relevant toxic oligomeric tau species.

No MeSH data available.


Related in: MedlinePlus

GSK-3β inhibition rescued microtubule number inhTau0N3RDrosophila, but increased total hTau protein and caused formation ofelectron-dense granules a-l) Electron micrographs of transverse sections ofperipheral nerves in L3 Drosophila (scale bar200 nm).In hTau-expressing (elavC155-Gal4/Y;UAS-hTau0N3R/+) animals treated with either20 mM LiCl (hTau-Li, a–c) or with20 μM AR-A01448 (hTau-AR, d-f), some axons exhibitedsmall electron-dense globular structures of approximately20–50 nm in size (black arrows). These structureswere extremely rare in control larvae expressingelavC155-Gal4 driver alone (WT,g–i) or untreatedhTau0N3R-expressing neurons (j–l).In WT larvae the axon profiles showed numerous regularly-spaced,correctly-aligned transverse microtubule profiles (black arrowheads in g-i;8.1 ± 0.2/axon profile). As we havepreviously shown29, in hTau0N3R-expressingaxons the microtubules were dramatically disrupted, with fewercorrectly-aligned transverse microtubule profiles (black arrowheads in j-l;5.3 ± 0.3/axon profile), and evidence ofdisorganised microtubules in the same axon profiles (white arrowheads inj–l). Indeed, approximately 30% ofhTau0N3R-expressing axons displayed no visiblemicrotubule profiles (FigureS1). In hTau0N3R-expressing larvae fed with Li(a–c) or AR (d–f),there were significantly more correctly-aligned transverse microtubuleprofiles (black arrowheads in a-l;9.2 ± 0.3/axon) and fewer misalignedmicrotubules. Microtubule numbers per axon are quantified in m(**p < 0.01, unpaired Students t test).Representative Western blots of hTau0N3R-expressing fly headlysates showed that tau phosphorylation was decreased (at T231/S235 detectedby AT180) whilst total tau levels were increased by 40–60%(o–r) by 20 mM lithium treatment(hTau-Li, o), 20 μM AR-A01448 treatment(hTau-AR, p), co-expression of dominant negative shaggy(hTau;sggDN, q){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sggDN/ + }.Conversely, total tau levels were decreased by approximately 50% byco-expression of constitutively active shaggy (hTau;sgg*, r){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sgg*/ + }. This isquantified in s (error bars are standard error of mean;*p < 0.05 by Students t-test).
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f1: GSK-3β inhibition rescued microtubule number inhTau0N3RDrosophila, but increased total hTau protein and caused formation ofelectron-dense granules a-l) Electron micrographs of transverse sections ofperipheral nerves in L3 Drosophila (scale bar200 nm).In hTau-expressing (elavC155-Gal4/Y;UAS-hTau0N3R/+) animals treated with either20 mM LiCl (hTau-Li, a–c) or with20 μM AR-A01448 (hTau-AR, d-f), some axons exhibitedsmall electron-dense globular structures of approximately20–50 nm in size (black arrows). These structureswere extremely rare in control larvae expressingelavC155-Gal4 driver alone (WT,g–i) or untreatedhTau0N3R-expressing neurons (j–l).In WT larvae the axon profiles showed numerous regularly-spaced,correctly-aligned transverse microtubule profiles (black arrowheads in g-i;8.1 ± 0.2/axon profile). As we havepreviously shown29, in hTau0N3R-expressingaxons the microtubules were dramatically disrupted, with fewercorrectly-aligned transverse microtubule profiles (black arrowheads in j-l;5.3 ± 0.3/axon profile), and evidence ofdisorganised microtubules in the same axon profiles (white arrowheads inj–l). Indeed, approximately 30% ofhTau0N3R-expressing axons displayed no visiblemicrotubule profiles (FigureS1). In hTau0N3R-expressing larvae fed with Li(a–c) or AR (d–f),there were significantly more correctly-aligned transverse microtubuleprofiles (black arrowheads in a-l;9.2 ± 0.3/axon) and fewer misalignedmicrotubules. Microtubule numbers per axon are quantified in m(**p < 0.01, unpaired Students t test).Representative Western blots of hTau0N3R-expressing fly headlysates showed that tau phosphorylation was decreased (at T231/S235 detectedby AT180) whilst total tau levels were increased by 40–60%(o–r) by 20 mM lithium treatment(hTau-Li, o), 20 μM AR-A01448 treatment(hTau-AR, p), co-expression of dominant negative shaggy(hTau;sggDN, q){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sggDN/ + }.Conversely, total tau levels were decreased by approximately 50% byco-expression of constitutively active shaggy (hTau;sgg*, r){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sgg*/ + }. This isquantified in s (error bars are standard error of mean;*p < 0.05 by Students t-test).

Mentions: We have previously shown that reduction of GSK3β-mediated tauphosphorylation (using LiCl or a more specific GSK-3β inhibitor,AR-A01448) rescues phenotypes induced by human tau (hTau0N3R) inDrosophila. These phenotypes include locomotor impairment anddisrupted axonal transport7893738. While examining theultrastructure of hTau0N3R-expressing neurons in these animals,we made an unexpected observation: treatment with either drug led to theformation of 20–50 nm electron-dense granules in axonalEM sections (Fig. 1, arrows in 1a-f and quantified in Supplementary Fig. 1). Thesestructures bear a striking resemblance to granular tau oligomers (GTOs) firstdescribed by the Takashima group in AD brains30. Subsequent invitro characterization by their group proved that AD brain GTOs weigh1800 kDa and contain on average 40 molecules of tau2439. Here, we tested the hypothesis that the electron-densegranules we observed in hTau-expressing Drosophila afterGSK-3β inhibition are indeed GTO-like structures.


Rescue from tau-induced neuronal dysfunction produces insoluble tau oligomers.

Cowan CM, Quraishe S, Hands S, Sealey M, Mahajan S, Allan DW, Mudher A - Sci Rep (2015)

GSK-3β inhibition rescued microtubule number inhTau0N3RDrosophila, but increased total hTau protein and caused formation ofelectron-dense granules a-l) Electron micrographs of transverse sections ofperipheral nerves in L3 Drosophila (scale bar200 nm).In hTau-expressing (elavC155-Gal4/Y;UAS-hTau0N3R/+) animals treated with either20 mM LiCl (hTau-Li, a–c) or with20 μM AR-A01448 (hTau-AR, d-f), some axons exhibitedsmall electron-dense globular structures of approximately20–50 nm in size (black arrows). These structureswere extremely rare in control larvae expressingelavC155-Gal4 driver alone (WT,g–i) or untreatedhTau0N3R-expressing neurons (j–l).In WT larvae the axon profiles showed numerous regularly-spaced,correctly-aligned transverse microtubule profiles (black arrowheads in g-i;8.1 ± 0.2/axon profile). As we havepreviously shown29, in hTau0N3R-expressingaxons the microtubules were dramatically disrupted, with fewercorrectly-aligned transverse microtubule profiles (black arrowheads in j-l;5.3 ± 0.3/axon profile), and evidence ofdisorganised microtubules in the same axon profiles (white arrowheads inj–l). Indeed, approximately 30% ofhTau0N3R-expressing axons displayed no visiblemicrotubule profiles (FigureS1). In hTau0N3R-expressing larvae fed with Li(a–c) or AR (d–f),there were significantly more correctly-aligned transverse microtubuleprofiles (black arrowheads in a-l;9.2 ± 0.3/axon) and fewer misalignedmicrotubules. Microtubule numbers per axon are quantified in m(**p < 0.01, unpaired Students t test).Representative Western blots of hTau0N3R-expressing fly headlysates showed that tau phosphorylation was decreased (at T231/S235 detectedby AT180) whilst total tau levels were increased by 40–60%(o–r) by 20 mM lithium treatment(hTau-Li, o), 20 μM AR-A01448 treatment(hTau-AR, p), co-expression of dominant negative shaggy(hTau;sggDN, q){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sggDN/ + }.Conversely, total tau levels were decreased by approximately 50% byco-expression of constitutively active shaggy (hTau;sgg*, r){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sgg*/ + }. This isquantified in s (error bars are standard error of mean;*p < 0.05 by Students t-test).
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f1: GSK-3β inhibition rescued microtubule number inhTau0N3RDrosophila, but increased total hTau protein and caused formation ofelectron-dense granules a-l) Electron micrographs of transverse sections ofperipheral nerves in L3 Drosophila (scale bar200 nm).In hTau-expressing (elavC155-Gal4/Y;UAS-hTau0N3R/+) animals treated with either20 mM LiCl (hTau-Li, a–c) or with20 μM AR-A01448 (hTau-AR, d-f), some axons exhibitedsmall electron-dense globular structures of approximately20–50 nm in size (black arrows). These structureswere extremely rare in control larvae expressingelavC155-Gal4 driver alone (WT,g–i) or untreatedhTau0N3R-expressing neurons (j–l).In WT larvae the axon profiles showed numerous regularly-spaced,correctly-aligned transverse microtubule profiles (black arrowheads in g-i;8.1 ± 0.2/axon profile). As we havepreviously shown29, in hTau0N3R-expressingaxons the microtubules were dramatically disrupted, with fewercorrectly-aligned transverse microtubule profiles (black arrowheads in j-l;5.3 ± 0.3/axon profile), and evidence ofdisorganised microtubules in the same axon profiles (white arrowheads inj–l). Indeed, approximately 30% ofhTau0N3R-expressing axons displayed no visiblemicrotubule profiles (FigureS1). In hTau0N3R-expressing larvae fed with Li(a–c) or AR (d–f),there were significantly more correctly-aligned transverse microtubuleprofiles (black arrowheads in a-l;9.2 ± 0.3/axon) and fewer misalignedmicrotubules. Microtubule numbers per axon are quantified in m(**p < 0.01, unpaired Students t test).Representative Western blots of hTau0N3R-expressing fly headlysates showed that tau phosphorylation was decreased (at T231/S235 detectedby AT180) whilst total tau levels were increased by 40–60%(o–r) by 20 mM lithium treatment(hTau-Li, o), 20 μM AR-A01448 treatment(hTau-AR, p), co-expression of dominant negative shaggy(hTau;sggDN, q){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sggDN/ + }.Conversely, total tau levels were decreased by approximately 50% byco-expression of constitutively active shaggy (hTau;sgg*, r){elavC155-Gal4/Y; UAS-hTau0N3R/ + ;UAS-sgg*/ + }. This isquantified in s (error bars are standard error of mean;*p < 0.05 by Students t-test).
Mentions: We have previously shown that reduction of GSK3β-mediated tauphosphorylation (using LiCl or a more specific GSK-3β inhibitor,AR-A01448) rescues phenotypes induced by human tau (hTau0N3R) inDrosophila. These phenotypes include locomotor impairment anddisrupted axonal transport7893738. While examining theultrastructure of hTau0N3R-expressing neurons in these animals,we made an unexpected observation: treatment with either drug led to theformation of 20–50 nm electron-dense granules in axonalEM sections (Fig. 1, arrows in 1a-f and quantified in Supplementary Fig. 1). Thesestructures bear a striking resemblance to granular tau oligomers (GTOs) firstdescribed by the Takashima group in AD brains30. Subsequent invitro characterization by their group proved that AD brain GTOs weigh1800 kDa and contain on average 40 molecules of tau2439. Here, we tested the hypothesis that the electron-densegranules we observed in hTau-expressing Drosophila afterGSK-3β inhibition are indeed GTO-like structures.

Bottom Line: Moreover, contrary to common belief, these tau oligomers were neither highly phosphorylated, and nor did they contain beta-pleated sheet structure.Whether these are inert or actively protective remains to be established.Nevertheless, this has wide implications for emerging therapeutic strategies such as those that target dissolution of tau oligomers as they may be ineffective or even counterproductive unless they act on the relevant toxic oligomeric tau species.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biological Sciences, University of Southampton, Southampton, SO17 1BJ, UK.

ABSTRACT
Aggregation of highly phosphorylated tau is a hallmark of Alzheimer's disease and other tauopathies. Nevertheless, animal models demonstrate that tau-mediated dysfunction/toxicity may not require large tau aggregates but instead may be caused by soluble hyper-phosphorylated tau or by small tau oligomers. Challenging this widely held view, we use multiple techniques to show that insoluble tau oligomers form in conditions where tau-mediated dysfunction is rescued in vivo. This shows that tau oligomers are not necessarily always toxic. Furthermore, their formation correlates with increased tau levels, caused intriguingly, by either pharmacological or genetic inhibition of tau kinase glycogen-synthase-kinase-3beta (GSK-3β). Moreover, contrary to common belief, these tau oligomers were neither highly phosphorylated, and nor did they contain beta-pleated sheet structure. This may explain their lack of toxicity. Our study makes the novel observation that tau also forms non-toxic insoluble oligomers in vivo in addition to toxic oligomers, which have been reported by others. Whether these are inert or actively protective remains to be established. Nevertheless, this has wide implications for emerging therapeutic strategies such as those that target dissolution of tau oligomers as they may be ineffective or even counterproductive unless they act on the relevant toxic oligomeric tau species.

No MeSH data available.


Related in: MedlinePlus