Browsing by Subject "Cryoelectron Microscopy"

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    Cryo-EM structures and functional characterization of homo- and heteropolymers of human ferritin variants
    (Nature, 2020-11-26) Irimia-Dominguez, Jose; Sun, Chen; Li, Kunpeng; Muhoberac, Barry B.; Hallinan, Grace I.; Garringer, Holly J.; Ghetti, Bernardino; Jiang, Wen; Vidal, Ruben; Pathology and Laboratory Medicine, School of Medicine
    The role of abnormal brain iron metabolism in neurodegenerative diseases is still insufficiently understood. Here, we investigate the molecular basis of the neurodegenerative disease hereditary ferritinopathy (HF), in which dysregulation of brain iron homeostasis is the primary cause of neurodegeneration. We mutagenized ferritin's three-fold pores (3FPs), i.e. the main entry route for iron, to investigate ferritin's iron management when iron must traverse the protein shell through the disrupted four-fold pores (4FPs) generated by mutations in the ferritin light chain (FtL) gene in HF. We assessed the structure and properties of ferritins using cryo-electron microscopy and a range of functional analyses in vitro. Loss of 3FP function did not alter ferritin structure but led to a decrease in protein solubility and iron storage. Abnormal 4FPs acted as alternate routes for iron entry and exit in the absence of functional 3FPs, further reducing ferritin iron-storage capacity. Importantly, even a small number of MtFtL subunits significantly compromises ferritin solubility and function, providing a rationale for the presence of ferritin aggregates in cell types expressing different levels of FtLs in patients with HF. These findings led us to discuss whether modifying pores could be used as a pharmacological target in HF.
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    Novel tau filament fold in corticobasal degeneration
    (Nature Publishing group, 2020-02-12) Zhang, Wenjuan; Tarutani, Airi; Newell, Kathy L.; Murzin, Alexey G.; Matsubara, Tomoyasu; Falcon, Benjamin; Vidal, Ruben; Garringer, Holly J.; Shi, Yang; Ikeuchi, Takeshi; Murayama, Shigeo; Ghetti, Bernardino; Hasegawa, Masato; Goedert, Michel; Scheres, Sjors H. W.; Pathology and Laboratory Medicine, School of Medicine
    Corticobasal degeneration (CBD) is a neurodegenerative tauopathy that is characterised by motor and cognitive disturbances (1–3). A higher frequency of the H1 haplotype of MAPT, the tau gene, is present in cases of CBD than in controls (4,5) and genome-wide association studies have identified additional risk factors (6). By histology, astrocytic plaques are diagnostic of CBD (7,8), as are detergent-insoluble tau fragments of 37 kDa by SDS-PAGE (9). Like progressive supranuclear palsy (PSP), globular glial tauopathy (GGT) and argyrophilic grain disease (AGD) (10), CBD is characterised by abundant filamentous tau inclusions that are made of isoforms with four microtubule-binding repeats (4R) (11–15). This distinguishes 4R tauopathies from Pick’s disease, filaments of which are made of three-repeat (3R) tau isoforms, and from Alzheimer’s disease and chronic traumatic encephalopathy (CTE), where both 3R and 4R tau isoforms are found in the filaments (16). Here we report the structures of tau filaments extracted from the brains of three individuals with CBD using electron cryo-microscopy (cryo-EM). They were identical between cases, but distinct from those of Alzheimer’s disease, Pick’s disease and CTE (17–19). The core of CBD filaments comprises residues K274-E380 of tau, spanning the last residue of R1, the whole of R2, R3 and R4, as well as 12 amino acids after R4. It adopts a novel four-layered fold, which encloses a large non-proteinaceous density. The latter is surrounded by the side chains of lysine residues 290 and 294 from R2 and 370 from the sequence after R4. CBD is the first 4R tauopathy with filaments of known structure.
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    Structures of filaments from Pick's disease reveal a novel tau protein fold
    (Nature Research, 2018-09) Falcon, Benjamin; Zhang, Wenjuan; Murzin, Alexey G.; Murshudov, Garib; Garringer, Holly J.; Vidal, Ruben; Crowther, R. Anthony; Ghetti, Bernardino; Scheres, Sjors H.W.; Goedert, Michel; Pathology and Laboratory Medicine, School of Medicine
    The ordered assembly of tau protein into abnormal filamentous inclusions underlies many human neurodegenerative diseases1. Tau assemblies seem to spread through specific neural networks in each disease2, with short filaments having the greatest seeding activity3. The abundance of tau inclusions strongly correlates with disease symptoms4. Six tau isoforms are expressed in the normal adult human brain-three isoforms with four microtubule-binding repeats each (4R tau) and three isoforms that lack the second repeat (3R tau)1. In various diseases, tau filaments can be composed of either 3R or 4R tau, or of both. Tau filaments have distinct cellular and neuroanatomical distributions5, with morphological and biochemical differences suggesting that they may be able to adopt disease-specific molecular conformations6,7. Such conformers may give rise to different neuropathological phenotypes8,9, reminiscent of prion strains10. However, the underlying structures are not known. Using electron cryo-microscopy, we recently reported the structures of tau filaments from patients with Alzheimer's disease, which contain both 3R and 4R tau11. Here we determine the structures of tau filaments from patients with Pick's disease, a neurodegenerative disorder characterized by frontotemporal dementia. The filaments consist of residues Lys254-Phe378 of 3R tau, which are folded differently from the tau filaments in Alzheimer's disease, establishing the existence of conformers of assembled tau. The observed tau fold in the filaments of patients with Pick's disease explains the selective incorporation of 3R tau in Pick bodies, and the differences in phosphorylation relative to the tau filaments of Alzheimer's disease. Our findings show how tau can adopt distinct folds in the human brain in different diseases, an essential step for understanding the formation and propagation of molecular conformers.