Cryo-EM structures of amyloid-ß and tau filaments in Down syndrome.

Adult individuals with Down syndrome (DS) develop Alzheimer disease (AD). Whether there is a difference between AD in DS and AD regarding the structure of amyloid-ß (Aß) and tau filaments is unknown. Here we report the structure of Aß and tau filaments from two DS brains. We found two Aß40 filaments (types IIIa and IIIb) that differ from those previously reported in sporadic AD and two types of Aß42 filaments (I and II) identical to those found in sporadic and familial AD. Tau filaments (paired helical filaments and straight filaments) were identical to those in AD, supporting the notion of a common mechanism through which amyloids trigger aggregation of tau. This knowledge is important for understanding AD in DS and assessing whether adults with DS could be included in AD clinical trials.

Cross-ß helical filaments of Tau and TMEM106B in Gray and White Matter of Multiple System Tauopathy with presenile Dementia.

Background: The Microtubule-Associated Protein Tau (MAPT) is one of the proteins that are central to neurodegenerative diseases. The nature of intracellular tau aggregates is determined by the cell types whether neuronal or glial, the participating tau isoforms, and the structure of the amyloid filament. The transmembrane protein 106B (TMEM106B) has recently emerged as another significant player in neurodegeneration and aging. In the central nervous system, the composition of the gray and white matter differs considerably. The gray matter consists of nerve cell bodies, dendrites, unmyelinated axons, synaptic terminals, astrocytes, oligodendrocytes (satellite cells) and microglia. The white matter differs from the gray for the presence of axonal tracts as the only neuronal component and for the absence of nerve cell bodies, dendrites and synaptic terminals. Cryogenic electron microscopy (cryo-EM) studies have unveiled the structure of tau and TMEM106B, from the cerebral cortex, in several neurodegenerative diseases; however, whether tau and TMEM106B filaments from the gray and white matter share a common fold requires additional investigation. Methods: We isolated tau and TMEM106B from the cerebral cortex and white matter of the frontal lobes of two individuals affected by multiple system tauopathy with presenile dementia (MSTD), a disease caused by the MAPT intron 10 mutation +3. We used immunostaining, biochemical, genetics and cryo-EM methods to characterize tau and TMEM106B. Results: We determined that tau filaments in the gray and the white matter of MSTD individuals can induce tau aggregation and have identical AGD type 2 folds. TMEM106B amyloid filaments were also found in the gray and white matter of MSTD; the filament folds were identical in the two anatomical regions. Conclusions: Our findings show for the first time that in MSTD two types of amyloid filaments extracted from the gray matter have identical folds to those extracted from the white matter. Whether in this genetic disorder there is a relationship in the pathogenesis of the tau and TMEM106B filaments, remains to be determined. Furthermore, additional studies are needed for other proteins and other neurodegenerative diseases to establish whether filaments extracted from the gray and white matter would have identical folds.

Cryo-EM structures of prion protein filaments from Gerstmann-Sträussler-Scheinker disease.

Prion protein (PrP) aggregation and formation of PrP amyloid (APrP) are central events in the pathogenesis of prion diseases. In the dominantly inherited prion protein amyloidosis known as Gerstmann-Sträussler-Scheinker (GSS) disease, plaques made of PrP amyloid are present throughout the brain. The c.593t > c mutation in the prion protein gene (PRNP) results in a phenylalanine to serine amino acid substitution at PrP residue 198 (F198S) and causes the most severe amyloidosis among GSS variants. It has been shown that neurodegeneration in this disease is associated with the presence of extracellular APrP plaques and neuronal intracytoplasmic Tau inclusions, that have been shown to contain paired helical filaments identical to those found in Alzheimer disease. Using cryogenic electron microscopy (cryo-EM), we determined for the first time the structures of filaments of human APrP, isolated post-mortem from the brain of two symptomatic PRNP F198S mutation carriers. We report that in GSS (F198S) APrP filaments are composed of dimeric, trimeric and tetrameric left-handed protofilaments with their protomers sharing a common protein fold. The protomers in the cross-ß spines consist of 62 amino acids and span from glycine 80 to phenylalanine 141, adopting a previously unseen spiral fold with a thicker outer layer and a thinner inner layer. Each protomer comprises nine short ß-strands, with the ß1 and ß8 strands, as well as the ß4 and ß9 strands, forming a steric zipper. The data obtained by cryo-EM provide insights into the structural complexity of the PrP filament in a dominantly inherited human PrP amyloidosis. The novel findings highlight the urgency of extending our knowledge of the filaments' structures that may underlie distinct clinical and pathologic phenotypes of human neurodegenerative diseases.

The effects of molecular crowding and CpG hypermethylation on DNA G-quadruplexes formed by the C9orf72 nucleotide repeat expansion.

A nucleotide repeat expansion (NRE), (G4C2)n, located in a classically noncoding region of C9orf72 (C9), is the most common genetic mutation associated with ALS/FTD. There is increasing evidence that nucleic acid structures formed by the C9-NRE may both contribute to ALS/FTD, and serve as therapeutic targets, but there is limited characterization of these nucleic acid structures under physiologically and disease relevant conditions. Here we show in vitro that the C9-NRE DNA can form both parallel and antiparallel DNA G-quadruplex (GQ) topological structures and that the structural preference of these DNA GQs can be dependent on the molecular crowding conditions. Additionally, 5-methylcytosine DNA hypermethylation, which is observed in the C9-NRE locus in some patients, has minimal effects on GQ topological preferences. Finally, molecular dynamic simulations of methylated and nonmethylated GQ structures support in vitro data showing that DNA GQ structures formed by the C9-NRE DNA are stable, with structural fluctuations limited to the cytosine-containing loop regions. These findings provide new insight into the structural polymorphic preferences and stability of DNA GQs formed by the C9-NRE in both the methylated and nonmethylated states, as well as reveal important features to guide the development of upstream therapeutic approaches to potentially attenuate C9-NRE-linked diseases.

Bending of the BST-2 coiled-coil during viral budding.

BST-2/tetherin is a human extracellular transmembrane protein that serves as a host defense factor against HIV-1 and other viruses by inhibiting viral spreading. Structurally, BST-2 is a homo-dimeric coiled-coil that is connected to the host cell membrane by N and C terminal transmembrane anchors. The C-terminal membrane anchor of BST-2 is inserted into the budding virus while the N-terminal membrane anchor remains in the host cell membrane creating a viral tether. The structural mechanism of viral budding and tethering as mediated by BST-2 is not clear. To more fully describe the mechanism of viral tethering, we created a model of BST-2 embedded in a membrane and used steered molecular dynamics to simulate the transition from the host cell membrane associated form to the cell-virus membrane bridging form. We observed that BST-2 did not transition as a rigid structure, but instead bent at positions with a reduced interface between the helices of the coiled-coil. The simulations for the human BST-2 were then compared with simulations on the mouse homolog, which has no apparent weak spots. We observed that the mouse homolog spread the bending across the ectodomain, rather than breaking at discrete points as observed with the human homolog. These simulations support previous biochemical and cellular work suggesting some flexibility in the coiled-coil is necessary for viral tethering, while also highlighting how subtle changes in protein sequence can influence the dynamics and stability of proteins with overall similar structure.