Cryo-EM structures of pathogenic fibrils and their impact on neurodegenerative disease research

Abstract

Neurodegenerative diseases are commonly associated with the formation of aberrant protein aggregates within the brain, and ultrastructural analyses have revealed that the proteins within these inclusions often assemble into amyloid filaments. Cryoelectron microscopy (cryo-EM) has emerged as an effective method for determining the near-atomic structure of these disease-associated filamentous proteins, and the resulting structures have revolutionized the way we think about aberrant protein aggregation and propagation during disease progression. These structures have also revealed that individual fibril conformations may dictate different disease conditions, and this newfound knowledge has improved disease modeling in the lab and advanced the ongoing pursuit of clinical tools capable of distinguishing and targeting different pathogenic entities within living patients. In this review, we summarize some of the recently developed cryo-EM structures of ex vivo α-synuclein, tau, β-amyloid (Aβ), TAR DNA-binding protein 43 (TDP-43), and transmembrane protein 106B (TMEM106B) fibrils and discuss how these structures are being leveraged toward mechanistic research and therapeutic development.

Keywords: Aβ; TDP-43; TMEM106B; amyloid; cryo-EM; tau; α-synuclein.

Figure 1:. The general structure of β-amyloid fibrils.

Amyloids, including neurodegenerative disease-associated fibrils, are filaments made up of one or more cross-β sheet protofilaments, which are comprised of stacked β-strands held together by hydrogen bonds.

Figure 2:. α-synuclein protofilament cores arise from its N-terminal and NAC domains.

(A) The full-length α-synuclein protein is depicted, with disease-specific protofilament cores shown as fragments. For all such molecular diagrams, numbers denote sequence residues. (B) Line drawings depict the filament cores of type I and type II MSA fibrils. Each fibril is a doublet with two different protofilament cores, one large and one small. Gray circle represents an unknown density at the protofilament-protofilament interface. N and C termini are marked. (C) An electrostatic surface representation and corresponding ribbon diagram of the Lewy fold is shown (PDB: 8A91). For this and all other electrostatic surface renderings, a color scheme is used to indicate the electrostatic potential: red is used to represent negative potential, white for neutral, and blue for positive potential. This coloring helps visualize the distribution of charges on the protein’s surface, which is important for understanding how the protein interacts with other molecules, such as ligands or cofactors. (D) The filament core region of the JOS fold is shown with the peptides that make up island A noted. Triangle indicates JOS insert. A line drawing shows a doublet fibril; singlets with an identical core fold are also observed. Gray circles represent unknown densities. N and C termini and islands A and B are marked on one protofilament.

Figure 3:. Tau fibrils map to the microtubule-binding domain of the protein and show a unique C-shaped conformation in AD PHFs.

(A) The full-length tau protein is depicted. 3R and 4R tau isoforms vary by the presence of the 1N and 2N insertions, and 3R isoforms lack R2. Protofilament cores from various tauopathies are shown as fragments; some recombinant tau fragments used in the lab are also included. (B) An electrostatic surface representation and ribbon model of the core of an AD PHF are shown (PDB: 5O3L).

Figure 4:. Different tauopathies are characterized by unique fibril core folds.

Protofilament cores from various tauopathies are depicted as line drawings with the N- and C-termini noted. Tauopathies are divided by the nature of the isoforms present in pathogenic deposits: AD and CTE accumulate both 3R and 4R isoforms, while Pick’s disease accumulates only 3R tau and other conditions accumulate only 4R tau. Except for AGD, doublets contain the same protofilament core fold as singlets. Gray circles represent unknown densities.

Figure 5:. Aβ(1-40) and Aβ(1-42) form ordered protofilament cores in AD.

(A) Full-length APP is cleaved by β- and γ-secretases to generate Aβ fragments, including Aβ(1-40) and Aβ(1-42). Aβ residues are numbered separately from that of APP. (B) An electrostatic surface representation and ribbon model are shown for a single Aβ(1-40) fibril (PDB: 6SHS). Each protofilament contains two Aβ(1-40) peptides. (C) The Aβ(1-42) ordered cores are shown as fragments and the overall shape of the fibril folds are shown as line drawings with N and C termini noted on one protofilament. Both polymorphs are doublets. (D) Aβ(1-42) ordered cores with the Arctic mutation (E22G, marked with asterisks) are shown as fragments. Fibrils are tetramers as shown, with two protofilaments in fold A (green) and two in fold B (blue). N and C termini noted on one protofilament per fold.

Figure 6:. TDP-43 protofilament cores map to the LCD of the protein.

(A) The full-length TDP-43 protein and disease-associated phosphorylation sites (P) are shown. The protofilament cores are shown as fragments. (B) An electrostatic surface representation of TDP-43 type A fibrils (PDB: 8CG3). (C) A ribbon model of the TDP-43 type A protofilament core is shown, while a line drawing represents the TDP-43 type B filament core fold. N and C termini are noted.

Figure 7:. The TMEM106B protofilament core maps to the luminal domain of the protein.

(A) The full-length TMEM106B protein and protofilament core fragment are depicted. TM: transmembrane domain. Asterisks mark residues 185 and 252. Glycosylation sites are marked with orange circles. (B) An electrostatic surface representation of a doublet filament with fold I is shown (PDB: 7SAS). (C) A ribbon model of a doublet with fold I is shown (top). Grey circle denotes the unidentified density at the protofilament-protofilament interface, asterisks mark residues 185 and 252, orange circles mark glycosylation sites. Line drawings (bottom) depict the other two protofilament core folds, including both variants of fold II. C termini are noted.