Biology subjects

Monistrol, J.

Publications and source records attributed to Monistrol, J..

6 recordsLinked to original sources

bioRxiv2025

Multi-immersion Oblique Plane Microscope (miOPM): A reconfigurable platform for high-resolution Light-Sheet Fluorescence Microscopy.

Light-Sheet Fluorescence Microscopy (LSFM) enables gentle, rapid, and efficient volumetric imaging of biological specimens. Despite its potential, LSFM has fragmented into multiple variants, each narrowly optimized for specific samples or imaging regimes, limiting its broad applicability. To overcome this barrier, we introduce the multi-immersion oblique plane microscope (miOPM), an adaptable LSFM platform for high-resolution imaging ranging from subcellular dynamics to whole organisms and cleared tissues. miOPM uniquely supports seamless interchangeability of oil, water, and air objectives, and maintains diffraction limited performance across a refractive index range of 1.33 - 1.51. It is compatible with standard sample mounting, current clearing protocols and high-throughput 3D imaging. We leverage miOPM to image diverse biological specimens at multiple spatial scales. By greatly expanding the adaptability and ease of use of LSFM, miOPM is poised to democratize advanced three-dimensional imaging.

bioengineering↗

bioRxiv2025

Structural and Functional Versatility of the Amyloidogenic Antimicrobial Peptide Citropin 1.3

Citropin 1.3 is an antimicrobial peptide produced by the amphibian Litoria citropa (Southern bell frog), which self-aggregates into distinct fibrillar structures, however, the function of the fibrils remains unclear and largely unexplored. In this study, the structural and functional properties of citropin 1.3 were investigated using cryogenic electron microscopy and fluorescence microscopy in the presence of membrane and cell models, and with X-ray crystallography. Canonical amyloids, multilayered nanotubes, and a novel mixed fibril were observed. Experiments with negatively charged giant unilamellar vesicles revealed that the peptide facilitates membrane fusion while simultaneously undergoing phase separation in the presence of phospholipids. In presence of mammalian cells, citropin 1.3 permeabilizes membranes, leading to cell death, and over time, colocalizes with genetic material. Overall, this work provides new insights into the structural dynamics of the amyloidogenic antimicrobial peptide citropin 1.3 and its interactions with different systems.

molecular biology↗

bioRxiv2024

Functional classification of tauopathy strains reveals the role of protofilament core residues.

Distinct tau amyloid assemblies underlie diverse tauopathies but defy rapid classification. Cell and animal experiments indicate tau functions as a prion, as different strains propagated in cells cause unique, transmissible neuropathology after inoculation. Strain amplification requires compatibility of the monomer and amyloid template. We used cryo-EM to study one cell-based YFP-tagged strain, resolving its amyloid nature. We then used sequential alanine (Ala) substitution (scan) within tau repeat domain (RD) to measure incorporation to pre-existing tau RD-YFP aggregates. This robustly discriminated strains, defining sequences critical for monomer incorporation. We then created 3R/4R or 4R WT RD (aa 246-408) biosensors. Ala scan of recombinant tau seeds with the Alzheimers Disease fold matched that of AD homogenate. We scanned 22 brain lysates comprising 4 tauopathies. This clustered cases by neuropathological syndrome, revealed the role of amino acids in protofilament folds, and allowed strain discrimination based on amino acid requirements for prion replication. TeaserDiscrimination of tau strains based on the relative contribution of each amino acid to templated propagation of the amyloid.

neuroscience↗

bioRxiv2024

Stepwise recruitment of Hsc70 by DNAJB1 produces ordered arrays primed for bursts of amyloid fibre disassembly

To understand the action of co-chaperones of the J-domain protein family in the assembly of disaggregation-active Hsc70 complexes on the surface of amyloid fibres, we used cryo-EM and tomography to compare the assemblies with wild-type co-chaperones or an activity deficient mutant. We show that human DNAJB1 binds uniformly and densely along -synuclein amyloid fibres in an asymmetric orientation, with one subunit of the DNAJB1 dimer lying along the fibre surface and the other subunit further away. It acts in a 2-step recruitment of Hsc70 to the fibres, first releasing DNAJB1 auto-inhibition and then activating the Hsc70 ATPase by the J domain, with ATPase recycling stimulated by the nucleotide exchange factor Apg2. When the auto-inhibition is removed by mutating the H5 inhibitory binding site on DNAJB1 ({Delta}1H5 DNAJB1 mutant), Hsc70 is recruited to the fibres at a normal level, but the resulting complex is inactive in disaggregation. Cryo tomography of the wild-type DNAJB1:Hsc70:Apg2:Syn fibre complex shows dense arrays of the chaperones extending out from the fibre surface along spiral tracks, with the Hsc70 density on the outer surface of the DNAJB1 layer. The {Delta}1H5 DNAJB1:Hsc70:Apg2:Syn fibre complex results in equally dense but less organised binding all over the fibre surface and lacks the ordered clusters. On the basis of these findings, we propose that 2-step activation of DNAJB1 regulates Hsc70 access to the fibre substrate. Since the DNAJB1 dimers are bound every 40 [A], the J domain released by Hsc70 binding to one dimer could activate the ATPase of an Hsc70 bound to the adjacent dimer. This could trigger a cascade of recruitment and activation in a localised, dense cloud of Hsc70 molecules to give coordinated, sequential binding and disaggregation from an exposed fibre end, as observed by cryo-EM and earlier fluorescence and AFM studies.

biochemistry↗

bioRxiv2023

Cryo-EM structure of Alzheimer disease tau filaments with PET ligand MK-6240

Positron Emission Tomography (PET) ligands have advanced Alzheimers disease (AD) diagnosis and treatment. Using autoradiography and cryo-EM, we identified AD brain tissue with elevated tau burden, purified filaments, and determined the structure of second-generation high avidity PET ligand MK-6240 at 2.31 [A] resolution, which bound at a 1:1 ratio within the cleft of tau paired-helical filament (PHF), engaging with glutamine 351, lysine K353, and isoleucine 360. This information elucidates the basis of MK-6240 PET in quantifying PHF deposits in AD and may facilitate the structure-based design of superior ligands against tau amyloids.

neuroscience↗

bioRxiv2021

Cooperative Amyloid Fibre Binding and Disassembly by the Hsp70 disaggregase

Although amyloid fibres are highly stable protein aggregates, a specific combination of human Hsp70 system chaperones can disassemble them, including fibres formed of -synuclein, huntingtin or Tau. Disaggregation requires the ATPase activity of the constitutively expressed Hsp70, Hsc70, together with the J domain protein DNAJB1 and the nucleotide exchange factor Apg2. Recruitment and clustering of Hsc70 on the fibrils appear to be necessary for disassembly. Here we use atomic force microscopy (AFM) to show that segments of in vitro assembled -synuclein fibrils are first coated with chaperones and then undergo bursts of rapid, unidirectional disassembly. Cryo-electron tomography reveals fibrils with regions of densely bound chaperones extending from the fibre surface, preferentially at one end of the fibre. Sub-stoichiometric amounts of Apg2 relative to Hsc70 dramatically increase recruitment of Hsc70 to the fibres, creating localised active zones that then undergo rapid disassembly at a rate of [~]4 subunits per second.

biochemistry↗