Cryo-EM reconstruction of helical polymers: Beyond the simple cases DOI
Mark A. B. Kreutzberger, Ravi R. Sonani, Edward H. Egelman

et al.

Quarterly Reviews of Biophysics, Journal Year: 2024, Volume and Issue: 57

Published: Jan. 1, 2024

Helices are one of the most frequently encountered symmetries in biological assemblies. Helical symmetry has been exploited electron microscopic studies as a limited number filament images, principle, can provide all information needed to do three-dimensional reconstruction polymer. Over past 25 years, reconstructions helical polymers from cryo-EM images have shifted completely Fourier-Bessel methods single-particle approaches. The approaches allowed people surmount problem that very few crystalline order, and despite flexibility heterogeneity present these polymers, reaching resolution where accurate atomic models be built now become standard. While determining correct may simple for something like F-actin, many other particularly those formed small peptides, it much more challenging. This review discusses why determination problematic, trial-and-error still best approach. Studies macromolecular assemblies, such icosahedral capsids, usually found not imposing leads great reduction while at same time revealing possibly interesting asymmetric features. We show certain assemblies sometimes lead greatly improved resolution. Further, case supercoiled flagellar filaments bacteria archaea, we imposition only wrong, but is necessary, obscures mechanisms whereby supercoil.

Language: Английский

Signalling by co-operative higher-order assembly formation: linking evidence at molecular and cellular levels DOI Creative Commons
Boštjan Kobe, Jeffrey D. Nanson, Mikayla Hoad

et al.

Biochemical Journal, Journal Year: 2025, Volume and Issue: 482(05), P. 275 - 294

Published: March 1, 2025

The concept of higher-order assembly signalling or by co-operative formation (SCAF) was proposed based on the structures assemblies formed proteins featuring domains from death-fold family and Toll/interleukin-1 receptor domain family. Because these form filamentous upon stimulation activate downstream pathways through induced proximity, they were envisioned to sharpen response thresholds extreme co-operativity assembly. Recent findings demonstrate that a central feature SCAF mechanism is nucleation barrier allows switch-like, digital 'all-or-none' minute stimuli. In agreement, this features in cell-death innate immunity activation where binary decision required. Here, we broaden encapsulate essential kinetic properties open-ended signalling, compare other such as biomolecular condensates, review how operates cells.

Language: Английский

Citations

1
Microscopy methods for the in vivo study of nanoscale nuclear organization DOI Creative Commons

Nidhi Rani Lokesh,

Mark E. Pownall

Biochemical Society Transactions, Journal Year: 2025, Volume and Issue: 53(01)

Published: Feb. 3, 2025

Eukaryotic genomes are highly compacted within the nucleus and organized into complex 3D structures across various genomic physical scales. Organization plays a key role in gene regulation, both facilitating regulatory interactions to promote transcription while also enabling silencing of other genes. Despite functional importance genome organization determining cell identity function, investigating nuclear this wide range scales has been challenging. Microscopy provides opportunity for direct visualization pioneered discoveries field. Nonetheless, nanoscale nucleus, such as nucleosomes chromatin loops, requires super-resolution imaging go beyond ~220 nm diffraction limit. Here, we review recent advances technology their promise uncover new insights at nanoscale. We discuss different modalities how they have applied with focus on light microscopy its application vivo systems. Finally, conclude our perspective continued technical innovations will advance understanding structure function.

Language: Английский

Citations

0
Conformational dynamics of the nuclear pore complex central channel DOI Creative Commons

Yu Chen,

Guoli Zhou,

Miao Yu

et al.

Biochemical Society Transactions, Journal Year: 2025, Volume and Issue: 53(01)

Published: Feb. 7, 2025

The nuclear pore complex (NPC) is a vital regulator of molecular transport between the nucleus and cytoplasm in eukaryotic cells. At heart NPC’s function are intrinsically disordered phenylalanineglycine-rich nucleoporins (FG-Nups), which form dynamic permeability barrier within central channel. This nature facilitates efficient nucleocytoplasmic but also poses significant challenges to its characterization, especially nano-confined environment NPC. Recent advances experimental techniques, such as cryo-electron microscopy, atomic force fluorescence magnetic resonance, along with computational modeling, have illuminated conformational flexibility FG-Nups, underpins their functional versatility. review synthesizes these advancements, emphasizing how disruptions FG-Nup behavior—caused by mutations or pathological interactions—contribute diseases neurodegenerative disorders, aging-related decline, viral infections. Despite progress, persist deciphering dynamics crowded cellular environment, under conditions. Addressing gaps critical for advancing therapeutic strategies targeting NPC dysfunction disease progression.

Language: Английский

Citations

0
SLICK: A Sandwich-LIke Culturing Kit for in situ Cryo-ET Sample Preparation DOI Creative Commons

Qiuye Li,

Li Zhang, Qin Xu

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2025, Volume and Issue: unknown

Published: Feb. 16, 2025

In situ cryo-electron tomography (cryo-ET) has recently been widely used in observing subcellular structures and macromolecules their native states at high resolution. One of the reasons that it not more adopted by cell biologists structural is difficulties sample preparation. Here we present Sandwich-LIke Culturing Kit (SLICK), simplifying procedure increasing throughput for preparation cryo-ET (69 words).

Language: Английский

Citations

0
Cryo-EM reconstruction of helical polymers: Beyond the simple cases DOI
Mark A. B. Kreutzberger, Ravi R. Sonani, Edward H. Egelman

et al.

Quarterly Reviews of Biophysics, Journal Year: 2024, Volume and Issue: 57

Published: Jan. 1, 2024

Helices are one of the most frequently encountered symmetries in biological assemblies. Helical symmetry has been exploited electron microscopic studies as a limited number filament images, principle, can provide all information needed to do three-dimensional reconstruction polymer. Over past 25 years, reconstructions helical polymers from cryo-EM images have shifted completely Fourier-Bessel methods single-particle approaches. The approaches allowed people surmount problem that very few crystalline order, and despite flexibility heterogeneity present these polymers, reaching resolution where accurate atomic models be built now become standard. While determining correct may simple for something like F-actin, many other particularly those formed small peptides, it much more challenging. This review discusses why determination problematic, trial-and-error still best approach. Studies macromolecular assemblies, such icosahedral capsids, usually found not imposing leads great reduction while at same time revealing possibly interesting asymmetric features. We show certain assemblies sometimes lead greatly improved resolution. Further, case supercoiled flagellar filaments bacteria archaea, we imposition only wrong, but is necessary, obscures mechanisms whereby supercoil.

Language: Английский

Citations

0