Paired Helical Filament Tau Research Articles

Association of Cancer History with Alzheimer's Disease Dementia and Neuropathology.

Cancer and Alzheimer's disease (AD) are common diseases of aging and share many risk factors. Surprisingly, however, epidemiologic data from several recent independent cohort studies suggest that there may be an inverse association between these diseases. To determine the relationship between history of cancer and odds of dementia proximate to death and neuropathological indices of AD. Using data from two separate clinical-pathologic cohort studies of aging and AD, the Religious Orders Study (ROS) and the Rush Memory and Aging Project (MAP), we compared odds of AD dementia proximate to death among participants with and without a history of cancer. We then examined the relation of history of cancer with measures of AD pathology at autopsy, i.e., paired helical filament tau (PHFtau) neurofibrillary tangles and amyloid-β load. Participants reporting a history of cancer had significantly lower odds of AD (OR 0.70 [0.55-0.89], p = 0.0040) proximate to death as compared to participants reporting no prior history of cancer. The results remained significant after adjusting for multiple risk factors including age, sex, race, education, and presence of an APOEɛ4 allele. At autopsy, participants with a history of cancer had significantly fewer PHFtau tangles (p < 0.001) than participants without a history of cancer, but similar levels of amyloid-β. Cancer survivors have reduced odds of developing AD and a lower burden of neurofibrillary tangle deposition.

Increased Caspase-6 activity in the human anterior olfactory nuclei of the olfactory bulb is associated with cognitive impairment.

Abnormally elevated hippocampal Caspase-6 (Casp6) activity is intimately associated with age-related cognitive impairment in humans and in mice. In humans, these high levels of Casp6 activity are initially localized in the entorhinal cortex, the area of the brain first affected by the formation of neurofibrillary tangles, according to Braak staging. The reason for the high vulnerability of entorhinal cortex neurons to neurofibrillary tangle pathology and Casp6 activity is unknown. Casp6 activity is involved in axonal degeneration, therefore, one possibility to explain increased vulnerability of the entorhinal cortex neurons would be that the afferent neurons of the olfactory bulb, some of which project their axons to the entorhinal cortex, are equally degenerating. To examine this possibility, we examined the presence of Casp6 activity, neurofibrillary tangle formation and amyloid deposition by immunohistochemistry with neoepitope antisera against the p20 subunit of active Casp6 and Tau cleaved by Casp6 (Tau∆Casp6), phosphorylated Tau paired helical filament (PHF-1) antibodies and anti-β-amyloid antiserum, respectively, in brains from individuals with no or mild cognitive impairment and Alzheimer disease (AD) dementia. Co-localization of Casp6 activity, PHF-1 and β-amyloid was detected mostly in the anterior olfactory nucleus (AON) of the olfactory bulb. The levels of active Casp6 in the AON, which were the highest in the AD brains, correlated with PHF-1 levels, but not with β-amyloid levels. AON Tau∆Casp6 levels correlated with entorhinal cortex Casp6 activity and PHF-1 levels. Multiple regression analyses demonstrated that AON Casp6 activity was associated with lower global cognitive function, mini mental state exam, episodic memory and semantic memory scores. These results suggest that AON Casp6 activity could lead to Casp6-mediated degeneration in the entorhinal cortex, but cannot exclude the possibilities that entorhinal cortex degeneration signals degeneration in the AON or that the pathologies occur in both regions independently. Nevertheless, AON Casp6 activity reflects that of the entorhinal cortex.

TAU BURDEN IS ASSOCIATED WITH SUBJECTIVE COGNITIVE CONCERNS IN THE CONTEXT OF β-AMYLOID BURDEN IN PRECLINICAL AD

Alzheimer’s disease (AD) is characterized by the deposition of β-amyloid (Aβ) and paired helical filament tau. The relationship of these deposits to the preclinical manifestations of the AD phenotype can now be evaluated in vivo with positron emission tomography (PET). Endorsement of subjective cognitive concerns (SCC) is associated with Aβ in this early phase of the disease trajectory, but until now, a relationship between SCC and regional tau deposition has not been interrogated. We aimed to elucidate the relationship between entorhinal cortical (EC) and inferior temporal (IT) tau, neocortical Aβ and SCC in clinically-normal (CN) older adults. Seventy-three CN (CDR = 0) participants (77yrs (SD=6.2), 58% female, 28% Aβ+, 28% APOEε4 carriers) participating in the Harvard Aging Brain Study underwent T807 (aka AV1451) and PiB PET imaging within 6 months of responding to three SCC questionnaires. An SCC composite was created using z-transformed memory subscales from the Memory Functioning Questionnaire, the Everyday Cognition Scale, and the adapted Structured Telephone Inventory for Dementia Assessment. Greater EC tau was associated with greater SCC (β=0.39, p<.001) after taking into account age, depressive symptoms, sex and education. IT tau was marginally associated (β=0.18, p=.16). After interactions between tau and Aβ were included, no interactive effect of EC tau and Aβ was found on SCC (p=.93), but an IT and Aβ interaction was now marginally associated with greater SCC (p=.07). This is the first study to demonstrate regionally-specific patterns of association between tau burden and SCC. Specifically, EC tau was strongly associated with greater SCC, but the effect was independent of Aβ burden. Increasing IT tau deposition, on the other hand, was related to greater SCC only within the context of increasing Ab burden. As such, both tau and Aβ are associated with SCC and appear to become multiplicative under certain circumstances. It is possible that IT tau-driven SCC reflects individuals likely on the AD trajectory, whereas EC-related SCC captures a range of phenomena including normal aging, in which EC tauopathy is very common. It remains to be seen whether regional tau underlies qualitatively different SCC phenotypes.

Tau PET: the next frontier in molecular imaging of dementia.

We have arrived at an exciting juncture in dementia research: the second major pathological hallmark of Alzheimer's disease (AD)-tau-can now be seen for the first time in the living human brain. The major proteinopathies in AD include amyloid-β plaques and neurofibrillary tangles (NFTs) made of hyperphosphorylated paired helical filament (PHF) tau. Since its advent more than a decade ago, amyloid PET imaging has revolutionized the field of dementia research, enabling more confident diagnosis of the likely pathology in patients with a variety of clinical dementia syndromes, paving the way for the identification of people with preclinical or prodromal AD pathology, and serving as a minimally invasive molecular readout in clinical trials of putative disease-modifying interventions. Now that we are on the brink of a second revolution in molecular imaging in dementia, it is worth considering the likely potential impact of this development on the field.

Tau pathology-dependent remodelling of cerebral arteries precedes Alzheimer's disease-related microvascular cerebral amyloid angiopathy.

Alzheimer’s disease (AD) is characterised by pathologic cerebrovascular remodelling. Whether this occurs already before disease onset, as may be indicated by early Braak tau-related cerebral hypoperfusion and blood–brain barrier (BBB) impairment found in previous studies, remains unknown. Therefore, we systematically quantified Braak tau stage- and cerebral amyloid angiopathy (CAA)-dependent alterations in the alpha-smooth muscle actin (α-SMA), collagen, and elastin content of leptomeningeal arterioles, small arteries, and medium-sized arteries surrounding the gyrus frontalis medialis (GFM) and hippocampus (HIPP), including the sulci, of 17 clinically and pathologically diagnosed AD subjects (Braak stage IV–VI) and 28 non-demented control subjects (Braak stage I–IV). GFM and HIPP paraffin sections were stained for general collagen and elastin with the Verhoeff–van Gieson stain; α-SMA and CAA/amyloid β (Aβ) were detected using immunohistochemistry. Significant arterial elastin degradation was observed from Braak stage III onward and correlated with Braak tau pathology (ρ = 0.909, 95 % CI 0.370 to 0.990, p < 0.05). This was accompanied by an increase in neutrophil elastase expression by α-SMA-positive cells in the vessel wall. Small and medium-sized arteries exhibited significant CAA-independent α-SMA loss starting between Braak stage I and II–III, along with accumulation of phosphorylated paired helical filament (PHF) tau in the perivascular space of intraparenchymal vessels. α-SMA remained at the decreased level throughout the later Braak stages. In contrast, arterioles exhibited significant α-SMA loss only at Braak stage V and VI/in AD subjects, which was CAA-dependent/correlated with CAA burden (ρ = −0.422, 95 % CI −0.557 to −0.265, p < 0.0001). Collagen content was only significantly changed in small arteries. Our data indicate that vessel wall remodelling of leptomeningeal arteries is an early-onset, Braak tau pathology-dependent process unrelated to CAA and AD, which potentially may contribute to downstream CAA-dependent microvascular pathology in AD.Electronic supplementary materialThe online version of this article (doi:10.1007/s00401-016-1560-2) contains supplementary material, which is available to authorized users.

Tau and neurodegenerative disease: the story so far.

In 1975, tau protein was isolated as a microtubule-associated factor from the porcine brain. In the previous year, a paired helical filament (PHF) protein had been identified in neurofibrillary tangles in the brains of individuals with Alzheimer disease (AD), but it was not until 1986 that the PHF protein and tau were discovered to be one and the same. In the AD brain, tau was found to be abnormally hyperphosphorylated, and it inhibited rather than promoted in vitro microtubule assembly. Almost 80 disease-causing exonic missense and intronic silent mutations in the tau gene have been found in familial cases of frontotemporal dementia but, to date, no such mutation has been found in AD. The first phase I clinical trial of an active tau immunization vaccine in patients with AD was recently completed. Assays for tau levels in cerebrospinal fluid and plasma are now available, and tau radiotracers for PET are under development. In this article, we provide an overview of the pivotal discoveries in the tau research field over the past 40 years. We also review the current status of the field, including disease mechanisms and therapeutic approaches.

Oral TNFα Modulation Alters Neutrophil Infiltration, Improves Cognition and Diminishes Tau and Amyloid Pathology in the 3xTgAD Mouse Model.

Cytokines such as TNFα can polarize microglia/macrophages into different neuroinflammatory types. Skewing of the phenotype towards a cytotoxic state is thought to impair phagocytosis and has been described in Alzheimer’s Disease (AD). Neuroinflammation can be perpetuated by a cycle of increasing cytokine production and maintenance of a polarized activation state that contributes to AD progression. In this study, 3xTgAD mice, age 6 months, were treated orally with 3 doses of the TNFα modulating compound isoindolin-1,3 dithione (IDT) for 10 months. We demonstrate that IDT is a TNFα modulating compound both in vitro and in vivo. Following long-term IDT administration, mice were assessed for learning & memory and tissue and serum were collected for analysis. Results demonstrate that IDT is safe for long-term treatment and significantly improves learning and memory in the 3xTgAD mouse model. IDT significantly reduced paired helical filament tau and fibrillar amyloid accumulation. Flow cytometry of brain cell populations revealed that IDT increased the infiltrating neutrophil population while reducing TNFα expression in this population. IDT is a safe and effective TNFα and innate immune system modulator. Thus small molecule, orally bioavailable modulators are promising therapeutics for Alzheimer’s disease.

Association of APOE with tau-tangle pathology with and without β-amyloid

This study tested the hypothesis that the association of apolipoprotein E (APOE) with paired helical filament tau (PHF-tau) tangle pathology differs in brains with and without β-amyloid. Participants were 1056 autopsied individuals from 2 clinical-pathologic cohort studies of aging and Alzheimer's disease (AD), the Religious Orders Study, and the Rush Memory and Aging Project. Neuropathologic measures were obtained using immunohistochemistry targeting β-amyloid and PHF-tau tangles in 8 brain regions. Linear regression was used to compare the relation of APOE ε4 and ε2 to PHF-tau-tangle density in persons with β-amyloid relative to persons without β-amyloid. We found an interaction between APOE ε4 carriers and presence of β-amyloid (β = −0.968, p = 0.013) such that the association of APOE ε4 with PHF-tau tangles was much stronger in brains with β-amyloid. Stratified analysis shows that the association of APOE ε4 with PHF-tau tangles was considerably stronger among those with β-amyloid (β = 0.757, p = 1.1 × 10−15) compared to those without β-amyloid which was not significant (β = −0.201, p = 0.424). Separately, APOE ε2 was associated with fewer tangles in brains with β-amyloid (β = −0.425, p = 7.6 × 10−4) compared to those without β-amyloid which was not significant (β = −0.102, p = 0.506). Thus, the presence of APOE ε4 and ε2 alleles was not associated with PHF-tau tangles in the absence of β-amyloid. The data provide additional evidence that PHF-tau tangles in the absence of β-amyloid may reflect a pathologic process distinct from Alzheimer's disease.

Association of Brain DNA methylation in SORL1, ABCA7, HLA-DRB5, SLC24A4, and BIN1 with pathological diagnosis of Alzheimer disease.

Recent large-scale genome-wide association studies have discovered several genetic variants associated with Alzheimer disease (AD); however, the extent to which DNA methylation in these AD loci contributes to the disease susceptibility remains unknown. To examine the association of brain DNA methylation in 28 reported AD loci with AD pathologies. Ongoing community-based clinical pathological cohort studies of aging and dementia (the Religious Orders Study and the Rush Memory and Aging Project) among 740 autopsied participants 66.0 to 108.3 years old. DNA methylation levels at individual CpG sites generated from dorsolateral prefrontal cortex tissue using a bead assay. Pathological diagnosis of AD by National Institute on Aging-Reagan criteria following a standard postmortem examination. Overall, 447 participants (60.4%) met the criteria for pathological diagnosis of AD. Brain DNA methylation in SORL1, ABCA7, HLA-DRB5, SLC24A4, and BIN1 was associated with pathological AD. The association was robustly retained after replacing the binary trait of pathological AD with 2 quantitative and molecular specific hallmarks of AD, namely, Aβ load and paired helical filament tau tangle density. Furthermore, RNA expression of transcripts of SORL1 and ABCA7 was associated with paired helical filament tau tangle density, and the expression of BIN1 was associated with Aβ load. Brain DNA methylation in multiple AD loci is associated with AD pathologies. The results provide further evidence that disruption of DNA methylation is involved in the pathological process of AD.

Assessing THK523 selectivity for tau deposits in Alzheimer’s disease and non–Alzheimer’s disease tauopathies

IntroductionThe introduction of tau imaging agents such as 18F-THK523 offers new hope for the in vivo assessment of tau deposition in tauopathies such as Alzheimer’s disease (AD), where preliminary 18F-THK523-PET studies have demonstrated significantly higher cortical retention of 18F-THK523 in AD compared to age-matched healthy individuals. In addition to AD, tau imaging with PET may also be of value in assessing non-AD tauopathies, such as corticobasal degeneration (CBD), progressive supranuclear palsy (PSP) and Pick’s disease (PiD).MethodsTo further investigate the ability of THK523 to recognize tau lesions, we undertook immunohistochemical and fluorescence studies in serial brain sections taken from individuals with AD (n = 3), CBD (n = 2), PSP (n = 1), PiD (n = 2) and Parkinson’s disease (PD; n = 2). In addition to the neuropathological analysis, one PSP patient had undergone a 18F-THK523 PET scan 5 months before death.ResultsAlthough THK523 labelled tau-containing lesions such as neurofibrillary tangles and neuropil threads in the hippocampus and frontal regions of AD brains, it failed to label tau-containing lesions in non-AD tauopathies. Furthermore, though THK523 faintly labelled dense-cored amyloid-β plaques in the AD frontal cortex, it failed to label α-synuclein-containing Lewy bodies in PD brain sections.ConclusionThe results of this study suggest that 18F-THK523 selectively binds to paired helical filament tau in AD brains but does not bind to tau lesions in non-AD tauopathies, or to α-synuclein in PD brains.

Suppressed Retinal Degeneration in Aged Wild Type and APPswe/PS1ΔE9 Mice by Bone Marrow Transplantation

Alzheimer's disease (AD) is an age-related condition characterized by accumulation of neurotoxic amyloid β peptides (Aβ) in brain and retina. Because bone marrow transplantation (BMT) results in decreased cerebral Aβ in experimental AD, we hypothesized that BMT would mitigate retinal neurotoxicity through decreased retinal Aβ. To test this, we performed BMT in APPswe/PS1ΔE9 double transgenic mice using green fluorescent protein expressing wild type (wt) mice as marrow donors. We first examined retinas from control, non-transplanted, aged AD mice and found a two-fold increase in microglia compared with wt mice, prominent inner retinal Aβ and paired helical filament-tau, and decreased retinal ganglion cell layer neurons. BMT resulted in near complete replacement of host retinal microglia with BMT-derived cells and normalized total AD retinal microglia to non-transplanted wt levels. Aβ and paired helical filament-tau were reduced (61.0% and 44.1% respectively) in BMT-recipient AD mice, which had 20.8% more retinal ganglion cell layer neurons than non-transplanted AD controls. Interestingly, aged wt BMT recipients also had significantly more neurons (25.4%) compared with non-transplanted aged wt controls. Quantitation of retinal ganglion cell layer neurons in young mice confirmed age-related retinal degeneration was mitigated by BMT. We found increased MHC class II expression in BMT-derived microglia and decreased oxidative damage in retinal ganglion cell layer neurons. Thus, BMT is neuroprotective in age-related as well as AD-related retinal degeneration, and may be a result of alterations in innate immune function and oxidative stress in BMT recipient mice.

Molecular Mechanism of Tau Aggregation Induced by Anionic and Cationic Dyes

Abnormal tau filaments are a hallmark of Alzheimer's disease. Anionic dyes such as Congo Red, Thiazine Red, and Thioflavin S are able to induce tau fibrillization in vitro. SH-SY5Y cells were incubated with each dye for seven days leading to intracellular aggregates of tau protein, with different morphological characteristics. Interestingly, these tau aggregates were not observed when the Methylene Blue dye was added to the cell culture. In order to investigate the molecular mechanisms underlying this phenomenon, we developed a computational model for the interaction of the tau paired helical filament (PHF) core with every dye by docking analysis. The polar/electrostatic and nonpolar contribution to the free binding energy in the tau PHF core-anionic dye interaction was determined. We found that the tau PHF core can generate a positive net charge within the binding site localized at residuesLys311 and Lys340 (numbering according to the longest isoform hTau40). These residues are important for the binding affinity of the negative charges present in the anionic dyes causing an electrostatic environment that stabilizes the complex. Tau PHF core protofibril-Congo Red interaction has a stronger binding affinity compared to Thiazine Red or Thioflavin S. By contrast, the cationic dye Methylene Blue does not bind to nor stabilize the tau PHF core protofibrils. These results characterize the driving forces responsible for the binding of tau to anionic dyes leading to their self-aggregation and suggest that Methylene Blue may act as a destabilizing agent of tau aggregates.

Cerebrovascular atherosclerosis correlates with Alzheimer pathology in neurodegenerative dementias

A growing body of evidence demonstrates an association between vascular risk factors and Alzheimer's disease. This study investigated the frequency and severity of atherosclerotic plaques in the circle of Willis in Alzheimer's disease and multiple other neurodegenerative diseases. Semi-quantitative data from gross and microscopic neuropathological examinations in 1000 cases were analysed, including 410 with a primary diagnosis of Alzheimer's disease, 230 with synucleinopathies, 157 with TDP-43 proteinopathies, 144 with tauopathies and 59 with normal ageing. More than 77% of subjects with Alzheimer's disease had grossly apparent circle of Willis atherosclerosis, a percentage that was significantly higher than normal (47%), or other neurodegenerative diseases (43-67%). Age- and sex-adjusted atherosclerosis ratings were highly correlated with neuritic plaque, paired helical filaments tau neurofibrillary tangle and cerebral amyloid angiopathy ratings in the whole sample and within individual groups. We found no associations between atherosclerosis ratings and α-synuclein or TDP-43 lesion ratings. The association between age-adjusted circle of Willis atherosclerosis and Alzheimer's disease-type pathology was more robust for female subjects than male subjects. These results provide further confirmation and specificity that vascular disease and Alzheimer's disease are interrelated and suggest that common aetiologic or reciprocally synergistic pathophysiological mechanisms promote both vascular pathology and plaque and tangle pathology.

The Synaptic Accumulation of Hyperphosphorylated Tau Oligomers in Alzheimer Disease Is Associated With Dysfunction of the Ubiquitin-Proteasome System

In Alzheimer disease (AD), deposition of neurofibrillary tangles and loss of synapses in the neocortex and limbic system each correlate strongly with cognitive impairment. Tangles are composed of misfolded hyperphosphorylated tau proteins; however, the link between tau abnormalities and synaptic dysfunction remains unclear. We examined the location of tau in control and AD cortices using biochemical and morphologic methods. We found that, in addition to its well-described axonal localization, normal tau is present at both presynaptic and postsynaptic terminals in control human brains. In AD, tau becomes hyperphosphorylated and misfolded at both presynaptic and postsynaptic terminals, and this abnormally posttranslationally modified tau is enriched in synaptoneurosomal fractions. Synaptic tau seems to be hyperphosphorylated and ubiquitinated, and forms stable oligomers resistant to SDS denaturation. The accumulation of hyperphosphorylated tau oligomers at human AD synapses is associated with increased ubiquitinated substrates and increased proteasome components, consistent with dysfunction of the ubiquitin-proteasome system. Our findings suggest that synaptic hyperphosphorylated tau oligomers may be an important mediator of the proteotoxicity that disrupts synapses in AD.

Human fetal tau protein isoform: Possibilities for Alzheimer's disease treatment

While early 1990s reports showed the phosphorylation pattern of fetal tau protein to be similar to that of tau in paired helical filaments (PHF) in Alzheimer's disease (AD), neither the molecular mechanisms of the transient developmental hyperphosphorylation of tau nor reactivation of the fetal plasticity due to re-expression of fetal protein kinases in the aging and AD human brain have been sufficiently investigated. Here, we summarize the current knowledge on fetal tau, adding new data on the specific patterns of tau protein and mRNA expression in the developing human brain as well as on change in tau phosphorylation in the perforant pathway after entorhinal cortex lesion in mice. As fetal tau isoform does not form PHF even in a highly phosphorylated state, understanding its expression and post-translational modifications represents an important avenue for future research towards the development of AD treatment and prevention.

Traumatic Brain Injury (TBI) Induces Down‐Regulation of Olfactory Receptors That Are Ectopically Expressed In The Brain: Implications In TBI‐Mediated Tauopathy

Traumatic brain injury (TBI) is a risk factor for Alzheimer's disease (AD). Tau‐related neuropathology is a key feature of both TBI and AD. We recently identified ectopic expression of selective olfactory receptors (OR) in peripheral blood mononuclear cells (PBMC) and in the brain. We demonstrated that reduced contents of certain ORs in PBMC may serve as biological surrogates (biomarkers) of TBI. Moreover, we found significantly reduced contents of OR biomarkers in post‐mortem brain specimens of cases with a history of TBI. Exploring the potential role of OR biomarkers in the regulation of abnormal tau processing, we found overexpression and activation of an OR biomarker in primary cortico‐hippocampal neurons significantly attenuates pathophysiological tau paired helical filament immunoreactivity. Thus, down regulation of certain OR biomarkers in the brain may causally promote pathophysiological mechanisms underlying TBI clinical complications and, possibly, increase risk of TBI patient to develop AD. Our evidence provides clinically accessible biomarkers for assessing the likelihood of TBI subjects to develop clinical complications, providing more sensitive outcome measurements for clinical interventions and, ultimately, the characterization of potential novel therapeutic targets to prevent TBI‐associated tauopathy.Supported by Department of Defense: Award W81XWH‐08‐2‐0026 to GMP

Some evolutionary perspectives on Alzheimer's disease pathogenesis and pathology

There is increasing urgency to develop effective prevention and treatment for Alzheimer's disease (AD) as the aging population swells. Yet, our understanding remains limited for the elemental pathophysiological mechanisms of AD dementia that may be causal, compensatory, or epiphenomenal. To this end, we consider AD and why it exists from the perspectives of natural selection, adaptation, genetic drift, and other evolutionary forces. We discuss the connection between the apolipoprotein E (APOE) allele and AD, with special consideration to APOE ɛ4 as the ancestral allele. The phylogeny of AD-like changes across species is also examined, and pathology and treatment implications of AD are discussed from the perspective of evolutionary medicine. In particular, amyloid-β (Aβ) neuritic plaques and paired helical filament tau (PHFtau) neurofibrillary tangles have been traditionally viewed as injurious pathologies to be targeted, but may be preservative or restorative processes that mitigate harmful neurodegenerative processes or may be epiphenoma of the essential processes that cause neurodegeneration. Thus, we raise fundamental questions about current strategies for AD prevention and therapeutics.

The Dynamic Structure of Filamentous Tau

Filaments of the protein tau are a characteristic occurrence in Alzheimer disease and many other neurodegenerative disorders and the distribution of tau filaments correlates well with the loss of neurons and cognitive functions in Alzheimer disease. Filament formation of tau filaments is based on structural transitions from random coil to b-structure, to give the paired helical filaments (PHFs) which share common characteristics of amyloid fibrils. Protease digestion and solvent-accessibility studies demonstrated that the “core” of PHFs is mainly built from repeat sequences in the C-terminal half of the tau protein. The PHF core is surrounded by a “fuzzy coat”, of more than 200 residues that come from the N-terminal half of the protein as well as the C-terminus (Figure 1a). Electron paramagnetic resonance and nuclear magnetic resonance (NMR) suggested that residues within the fuzzy coat are highly flexible. Biochemical studies have shown that the fuzzy coat is important for tau aggregation as well as neurotoxicity. Herein we characterized the dynamic structure of PHFs formed by 441-residue tau (htau40), the longest isoform of tau present in the human central nervous system (Figure 1a), at single-residue level using NMR spectroscopy. We aggregated N-labeled htau40 into insoluble filaments. NMR diffusion experiments demonstrated that the observed NMR signals arises from aggregated tau protein with a molecular mass of more than 1 MDa (Figure 1b). In a two-dimensional heteronuclear single quantum coherence (HSQC) spectrum employing high-resolution magic-angle spinning (HR-MAS) (see Figure S1 in the Supporting Information), we observed about 260 signals (Figure 1c and Figure S2 and S3 in the Supporting Information). Sequencespecific resonance assignment of 244 of these signals (BMRB accession number: 17920; see Figure S2 in the Supporting Information) identified most of the residues in the N-terminal domain up to Thr212 and at the C-terminus starting at Val399. No signals were detected for residues between Thr212 and Val399, suggesting that residues in the central domain are too immobile to be detected by liquid-state NMR spectroscopy in agreement with previous studies. Comparison with monomeric htau40 revealed that the NMR resonances of many residues were strongly reduced in filamentous htau40 (Figure 1d,e). Most strikingly, the sections His121– Lys130 and Met1–Gly37 that are separated from the fibril core by 170 residues or more, showed changes in position and intensity of NMR signals (Figure 1e and Figure S2c in the Supporting Information). The presence of chemical exchange in these regions was further supported by N spin relaxation measurements (Figure 1 f and Supporting Figure S3). In agreement with chemical exchange, additional peaks were observed in close proximity to several of these residues (Figure 1 f and Figure S4 in the Supporting Information). The additional signals could not be connected in triple-resonance experiments or using exchange spectroscopy because of low signal-to-noise and signal overlap. Therefore, we assigned the additional peaks to the residue for which the assigned crosspeak of the major peak set had the greatest similarity in chemical shifts and paramagnetic relaxation enhancement (see Figure S5 in the Supporting Information). This procedure indicates that the additional peaks arise from residues at the Nand C-terminus. No peak doubling was observed at the Nand C-terminus in monomeric tau (see Figure S4 in the Supporting Information), highlighting the specificity of the multiple conformations in PHF tau. We revealed the identity of the PHF-specific conformations through measurements of paramagnetic relaxation enhancements (PREs), in which nitroxide spin labels are attached to cysteine residues at various positions in the PHF tau. The resulting broadening of amide resonances caused by enhanced relaxations rate through the paramagnetic nitroxide label, is quantified through the intensity ratios in the paramagnetic and diamagnetic states (Figure 2). The PRE effect scales as the inverse sixth power of the distance between the unpaired electron of the nitroxide unit and the NMR spin, providing a powerful probe of distances. N spin relaxation times (Figure 1 f) indicate that the fuzzy coat of PHFs is highly dynamic on a broad scale suggesting that the correlation time of the electron—amide proton internuclear vector is comparable to that of small water soluble proteins. Initially, we measured PRE broadening for PHFs with a [*] S. Bibow, Dr. M. D. Mukrasch, Prof. Dr. C. Griesinger, Prof. Dr. M. Zweckstetter Department of NMR-based Structural Biology, Max Planck Institute for Biophysical Chemistry Am Fassberg 11, 37077 Gcttingen (Germany) E-mail: mzwecks@gwdg.de

Accumulation of Transactive Response DNA Binding Protein 43 in Mild Cognitive Impairment and Alzheimer Disease

Transactive response DNA binding protein 43 (TDP-43) plays a central role in the neuropathology of frontotemporal lobar degeneration and amyotrophic lateral sclerosis, but the relationship between TDP-43 abnormalities and Alzheimer disease (AD) remains unclear. To determine whether TDP-43 can serve as a neuropathologic marker of AD, we performed biochemical characterization and quantification of TDP-43 in homogenates from parietal neocortex of subjects with aclinical diagnosis of no cognitive impairment (NCI, n = 12), mild cognitive impairment (MCI, n = 12), or AD (n = 12). Immunoblots revealed increased detergent-insoluble TDP-43 in the cortex of 0, 3, and 6 of the 12 individuals with NCI, MCI, or AD, respectively. Detergent-insoluble TDP-43 was positively correlated with the accumulation of soluble Aβ42, amyloid plaques, and paired helical filamenttau. In contrast, phospho-TDP-43 was decreased in the cytosolic fraction and detergent-soluble membrane/nuclear fraction from AD patients and correlated with antemortem cognitive function.Immunofluorescence analysis confirmed that the frequencies of individuals with TDP-43 or phospho-TDP-43 cytoplasmic inclusions were higher in AD than in NCI, with MCI at an intermediate level. These data indicate that abnormalities of TDP-43 occur in an important subset of MCI and AD patients and that they correlate with the clinical and neuropathologic features of AD.

Foxg1 coordinates the timing of projection neuron production in the cerebral cortex

This study examined the phosphorylation of tau on Ser 262, within the first microtubule-binding domain, by a developmentally regulated 100 kDa protein kinase exhibiting significantly greater activity in the embryonic rat brain than in the adult rat brain. This protein kinase co-purified with microtubules and co-immunoprecipitated with both tau and MAP-2. In addition to phosphorylating tau, MAP-2, and a Ser 262-containing peptide, the present protein kinase activity was shown to autophosphorylate as determined by the in-gel kinase assay in the absence of any protein or peptide polymerized into the matrix. Phosphorylation of tau with this protein kinase significantly reduced the tau–microtubule interaction, and the effect was significantly greater with microtubule-associated protein (MAP) preparations from embryonic brain than with preparations from the adult. Ser 262 is phosphorylated extensively in paired helical filament (PHF) tau from Alzheimer's disease (AD) brain, to a lesser extent in fetal tau, and only to a very minor extent in biopsy-derived human tau. Because the 100 kDa protein kinase activity phosphorylates Ser 262 and is higher in the fetal brain than the adult brain, it is hypothesized that an inappropriate re-expression and/or re-activation of this or a similar developmentally regulated protein kinase could contribute to the phosphorylation of Ser 262 in PHF-tau, and thus play a role in the pathogenesis of AD.