In Alzheimer disease research, focus has recently shifted away from large plaques and toward small oligomers of amyloid as a potential cause of disease (e.g., ARF related news story on Pigino et al., 2009 and Moreno et al., 2009). At the Society for Neuroscience annual meeting held 17-21 October 2009 in Chicago, Illinois, dozens of presentations dealt with Aβ oligomers, but what stood out more as a budding trend were signs that research in the tau and α-synuclein fields may be going down the same road. Several presentations covered emerging tools to study oligomers of these two proteins, and they drew wide notice among attendees.

Untangling Tau
Oligomers form shapes distinct from either monomer or fibrillar aggregated forms. Accordingly, researchers can make antibodies specific for particular conformations. Rakez Kayed described how his group at the University of Texas Medical Branch in Galveston pursued a similar approach to generate first tau oligomers, then antibodies against them, that had borne fruit while he was a postdoc in Charlie Glabe’s laboratory at the University of California, Irvine (Kayed et al., 2003). In his Galveston lab, Kayed seeded recombinant full-length human tau with Aβ oligomers or with α-synuclein oligomers. “I think this way of making oligomers is relevant to what happens in vivo,” Kayed told ARF. The scientists used the resulting tau oligomers as an antigen and produced a series of anti-tau antibodies that they claim recognize trimers and higher-order oligomers, but not monomers or fibrils.

The lab gave several presentations in Chicago. In a slide talk, Kayed focused on tau oligomers with a distinct ring-like shape. The story starts with work on amyloid-β, which can form distinct pore-like structures called annular protofibrils. As postdoc, Kayed had raised an antibody, dubbed Officer, specific for these shapes. He and Glabe found that Officer also nabbed other pore-forming proteins, suggesting it is specific not for amyloid-β, but for the β-barrel structural motif of the pore (Kayed et al., 2009). In Chicago, Kayed described how when he used Officer to detect amyloid-β annular protofibrils in brain tissue from people who had had Alzheimer disease, he noticed that some of the labeled annular structures contained no amyloid-β and looked suspiciously like structures commonly formed by tau. Sure enough, double-immunostaining with tau antibodies and Officer suggested that the AD brain contains tau, as well as amyloid-β, annular protofibrils. The scientists found similar tau annular protofibrils in tissue from people who had dementia with Lewy bodies (DLB) and a tauopathy called progressive supranuclear palsy (see also ARF related news story). The researchers apparently were also able to make Officer-reactive tau oligomers in vitro; on a Western blot, these tau annular protofibrils ran as a smear with a molecular weight more than 50 kDa, Kayed reported.

Kayed suggested that tau annular protofibrils might form pores in the cell membrane, allowing ions to pass through and disrupting cellular homeostasis. This might be true especially in astrocytes and oligodendrocytes, which contain more annular tau structures than do neurons, Kayed said. Because the annular protofibrils are ubiquitinated, they might also impede the cell’s protein destruction pathway, he added, allowing other misfolded proteins to accumulate. For background on the pore hypothesis, see ARF Live Discussion.

On a poster, Cristian Lasagna-Reeves, a graduate student in Kayed’s laboratory, presented a similar approach with a different antibody that detects tau oligomers. Lasagna-Reeves noted that the presence of neurofibrillary tangles correlates less tightly with symptoms of AD than does the presence of pre-fibrillar tau. “We think [tau oligomers] are the real toxic species in Alzheimer disease,” Lasagna-Reeves said, adding that his work now gives researchers a method to detect those species.

Called T2286, his new antibody detects 109-kDa spherical oligomers, but not monomers or fibrils, nor other kinds of amyloid-forming protein. Using brain tissue extracts, Lasagna-Reeves reported that frontal cortex from people who had had AD showed immunoreactivity with T2286; control samples did not. Using CSF from some 20 people, he reported an increase in T2286 immunoreactivity in people with AD relative to normal controls, suggesting spherical tau could eventually become a biomarker for the disease as well as a potential therapeutic target. This early CSF data was not presented alongside established CSF tau assays, for example, the INNOTEST htau or phospho-tau ELISAs widely used in clinical AD research across the world, and no absolute concentrations of the tau oligomers in CSF were given on the poster. Therefore, a comparison of the proposed tau oligomers to prior data on CSF total tau or phospho-tau concentrations in AD and controls was not possible from this initial study. In the next few months, the Texas group is planning to measure tau oligomers in a large number of CSF and serum samples, Kayed noted.

In a conversation with ARF, Kayed said his lab did do a comparison both in CSF and brain extracts, that is, with various research antibodies against phosphorylated forms of tau. It showed that less than 30 percent of the tau oligomers his new antibody detected were phosphorylated. “This surprised us. We had expected it would be more,” Kayed said. Whether phosphorylated tau might act as a seed, or whether phosphorylation might not occur until after filaments have formed, remains debatable, Kayed noted. Beyond AD brain, the T2286 antibody detected tau oligomers in brain extracts of dementia with Lewy bodies, supranuclear palsy, and Parkinson disease (see ARF related news story). In addition, T2286 picked up tau oligomers in the P301L and rTg4510 tau mouse models, as well as an α-synuclein mouse model, the Tg2576, and an APP/PS1 line, Kayed said.

In the rTg4510 regulatable tau mouse, the presence of oligomers correlated with a behavioral phenotype Karen Ashe’s group at the University of Minnesota, Minneapolis, had demonstrated previously, whereby memory function recovers when the tau transgene gets switched off even as neurofibrillary tangles stay in place (Santacruz et al., 2005; Berger et al., 2007). “Our new contribution to that is showing specifically with our antibody that the oligomers are gone at this point of behavior improvement,” Kayed said. He added that other unpublished experiments indicate that these oligomers are toxic to cultured cells, and that scientists at University of Texas Medical Branch plan to begin humanizing this antibody toward a tau immunotherapy (see also Kayed and Jackson, 2009) and to elucidate the epitope recognized by this antibody.

Overall, other scientists were intrigued by this talk and poster. However, they cautioned that the prospect of a single antibody that specifically recognizes neurotoxic tau oligomers in Western blots, ELISA, on tissue sections, in CSF, and that can serve as a start for tau oligomer immunotherapy sounds almost too good to be true and will need careful experimental substantiation.

In another poster, Kristina Patterson, a graduate student in the laboratory of Skip Binder at the Northwestern University School of Medicine in Chicago, presented her work on tau oligomers. Patterson used chemical cross-linking to stabilize tau oligomers in vitro. Typical cross-linkers rely on tau’s two cysteines to hook peptides together, but that limits the linkable conformations to those with two cysteines in proximity. “We decided we wanted to give it more choices,” Patterson said. She used a benzophenone cross-linker that binds cysteine with one end, but any carbon-hydrogen bond with the other. The non-specific end is activated by ultraviolet light. In other words, Patterson allowed the cross-linker to bind the cysteines and then gave the tau molecules the opportunity to oligomerize before hitting them with UV light to stabilize whatever conformation they happened to be in.

Patterson rather expected to see a ladder in her gels, with step-like increases for each additional tau in the oligomer. Instead, she saw mostly a species that ran at 180 kDa. The researchers also discovered a 180-kDa tau oligomer in brain homogenates from four people who had AD; the oligomer was not present in control samples. The 180 kDa corresponds to a trimer, but Patterson noted that other tau combinations could run at that weight depending on their structure. Patterson is currently characterizing the exact makeup of the oligomers, but has not yet performed size exclusion chromatography to show definitively that the bands from the AD sample are oligomers. In general, the study of tau oligomers is in its infancy, Patterson said, opening up many new possibilities for research. “These results overall are similar to what we see,” Kayed told ARF at the conference.

Un-aggregating α-Synuclein
Test-tube cross-linkers are also contributing to α-synuclein research. Martin Ingelsson presented a talk on work he and Joakim Bergström are doing at Uppsala University in Sweden. They used reactive aldehydes; these are compounds formed in the body under conditions of oxidative stress, a likely contributor to α-synucleinopathies. The researchers compared α-synuclein oligomers cross-linked by either 4-hydroxy-2-nonenal (HNE) or 4-oxo-2-nonenal (ONE). Both of these reactive aldehydes converted α-synuclein monomers into stable, β-sheet rich structures of approximately 2,000 kDa, which Ingelsson estimates contain 40 to 50 monomers. However, atomic force microscopy revealed that the ONE-induced α-synuclein oligomers were amorphous and variable in size, while HNE-induced oligomers formed distinct donut-shaped structures that are similar to the annular protofibrils described for amyloid-β and tau. None of these oligomers aggregated further into fibrils such as seen in Lewy bodies. This supports a suggestion made previously by Glabe, Paul Muchowski, and others that oligomers can form in their own side pathways that dead-end with the oligomeric state and do not continue on to large fibrils.

Ingelsson further reported that both types of oligomer were taken up by cultured cells, where they proved cytotoxic (see also Vekrellis data in ARF related news story). “This is just another example of commonalities” in diseases based on amyloid-forming proteins, Ingelsson said. Based on this research, the Uppsala group is hoping to develop a future α-synuclein immunotherapy along the lines of an anti-Aβ oligomer antibody originally developed in their lab, which is in late preclinical development at present.

Last but not least, Karin Danzer, a postdoctoral researcher in the laboratory of Pam McLean at Massachusetts General Hospital in Charlestown, presented new data on how she detected α-synuclein oligomers secreted from living cells. The poster generated a crowd and persistent buzz among scientists. “This demonstration of α-synuclein oligomer secretion is very attractive, because it implies these species will be more accessible to therapeutic removal than previously thought,” commented Dennis Selkoe of Brigham and Women’s Hospital in Boston. Danzer characterized the transmission of oligomers through cell media. The group combined culturing neuroglioma cells in a microfluidic chamber with an assay they developed to determine if extracellular α-synuclein was monomeric or oligomeric. Danzer generated two α-synuclein fusions: one linking the protein to the amino terminus of luciferase, and one linking it to the carboxyl terminus. Alone, the fusion proteins produced no luminescence. Together in an oligomer, the luciferase domains worked together to release light, which Danzer measured to quantify oligomerization.

The researchers found that α-synuclein oligomers, ranging in size from 14 kDA to 40,000 kDa, were secreted by neuroglioma cells. In previous work, the same group had shown that the chaperone Hsp70 reduced α-synuclein aggregation (Klucken et al., 2004). In the new assay, they found that co-transfecting Hsp70 reduced the secreted oligomer signal by 24-fold, without decreasing the amount of α-synuclein present. The data suggest that Hsp70 blocks oligomerization without destroying the protein. This presentation was also summarized on PD Online).—Amber Dance and Gabrielle Strobel.


No Available Comments

Make a Comment

To make a comment you must login or register.


News Citations

  1. The Many Misdeeds of Aβ—Seizures and Axonal Transport Interference
  2. Like DLB, Like AD—Do Oligomers Stir Up the Trouble?
  3. Et tu, Brute? Parkinson’s GWAS Fingers Tau Next to α-Synuclein
  4. Still Early Days for α-synuclein Fluid Marker

Webinar Citations

  1. Now You See Them, Now You Don't: The Amyloid Channel Hypothesis

Paper Citations

  1. . Disruption of fast axonal transport is a pathogenic mechanism for intraneuronal amyloid beta. Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5907-12. PubMed.
  2. . Synaptic transmission block by presynaptic injection of oligomeric amyloid beta. Proc Natl Acad Sci U S A. 2009 Apr 7;106(14):5901-6. PubMed.
  3. . Common structure of soluble amyloid oligomers implies common mechanism of pathogenesis. Science. 2003 Apr 18;300(5618):486-9. PubMed.
  4. . Annular protofibrils are a structurally and functionally distinct type of amyloid oligomer. J Biol Chem. 2009 Feb 13;284(7):4230-7. PubMed.
  5. . Tau suppression in a neurodegenerative mouse model improves memory function. Science. 2005 Jul 15;309(5733):476-81. PubMed.
  6. . Accumulation of pathological tau species and memory loss in a conditional model of tauopathy. J Neurosci. 2007 Apr 4;27(14):3650-62. PubMed.
  7. . Prefilament tau species as potential targets for immunotherapy for Alzheimer disease and related disorders. Curr Opin Immunol. 2009 Jun;21(3):359-63. PubMed.
  8. . Hsp70 Reduces alpha-Synuclein Aggregation and Toxicity. J Biol Chem. 2004 Jun 11;279(24):25497-502. PubMed.

Other Citations

  1. rTg4510

External Citations

  1. P301L
  2. PD Online

Further Reading


  1. . Inclusion formation and neuronal cell death through neuron-to-neuron transmission of alpha-synuclein. Proc Natl Acad Sci U S A. 2009 Aug 4;106(31):13010-5. PubMed.
  2. . Seeding induced by alpha-synuclein oligomers provides evidence for spreading of alpha-synuclein pathology. J Neurochem. 2009 Oct;111(1):192-203. PubMed.
  3. . Single particle characterization of iron-induced pore-forming alpha-synuclein oligomers. J Biol Chem. 2008 Apr 18;283(16):10992-1003. PubMed.
  4. . Functional protein kinase arrays reveal inhibition of p-21-activated kinase 4 by alpha-synuclein oligomers. J Neurochem. 2007 Dec;103(6):2401-7. PubMed.
  5. . Different species of alpha-synuclein oligomers induce calcium influx and seeding. J Neurosci. 2007 Aug 22;27(34):9220-32. PubMed.
  6. . The impact of the E46K mutation on the properties of alpha-synuclein in its monomeric and oligomeric states. Biochemistry. 2007 Jun 19;46(24):7107-18. PubMed.
  7. . Are amyloid diseases caused by protein aggregates that mimic bacterial pore-forming toxins?. Q Rev Biophys. 2006 May;39(2):167-201. PubMed.
  8. . Common structure and toxic function of amyloid oligomers implies a common mechanism of pathogenesis. Neurology. 2006 Jan 24;66(2 Suppl 1):S74-8. PubMed.
  9. . Amyloid ion channels: a common structural link for protein-misfolding disease. Proc Natl Acad Sci U S A. 2005 Jul 26;102(30):10427-32. PubMed.
  10. . The amyloid hypothesis of Alzheimer's disease: progress and problems on the road to therapeutics. Science. 2002 Jul 19;297(5580):353-6. PubMed.
  11. . Neurodegenerative disease: amyloid pores from pathogenic mutations. Nature. 2002 Jul 18;418(6895):291. PubMed.
  12. . Inherent toxicity of aggregates implies a common mechanism for protein misfolding diseases. Nature. 2002 Apr 4;416(6880):507-11. PubMed.