Journal: Acta Neuropathologica Communications
Article Title: Mimicking hypomethylation of FUS requires liquid–liquid phase separation to induce synaptic dysfunctions
doi: 10.1186/s40478-023-01703-w
Figure Lengend Snippet: FUS-16R causes the formation of dendritic inclusions which exhibit spontaneous movement that is enhanced by neuronal activity . A Representative confocal images illustrating the somatic and dendritic region of CA1 neurons expressing either FUS-16R-EYFP (FUS-16R) and td-tomato (top panels; cell 1–4) or FUS-WT-EYFP (FUS-WT) and td-tomato (bottom panels; cell 5–8). B Representative straightened time-lapse image of an apical dendritic region transfected with FUS-16R top panel (1 s), middle panel (6 s). Live imaging allowed for the tracking of individual FUS-16R condensates as indicated by the pseudo-colour merged time-lapse image (bottom panel; green 1 s; magenta 6 s). C Quantification of the FUS-16R-EYFP granule trajectory length recorded from individual granules. D Representative pseudo-coloured heat map of FUS16R condensate intensity in a dendritic region of interest. Dendritic FUS16R condensates are highlighted (ROI1-3) and their movement trajectory over a 1-min period is plotted for pre-stimulation (baseline; black line) and post-stimulation (Chrismon depolarisation; red line). E Quantification of normalised FUS-16R-condensate movement (averaged 5–6 ROIs per cell, n = 6). Average trajectory length was longer post-stimulation (p = 0.009373, paired t-test). F Representative multiphoton timelapse (10-min interval) heat maps of FUS-16R intensity at a single CA1 dendritic spine prior to and following single spine glutamate uncaging. A single spine was stimulated (cyan dot) and the FUS-16R intensity was measured by line scan across spine head. Histogram (below) illustrating FUS-16R condensate signal following stimulation in the presence (red bars) and absences (grey bars) of MNI-glutamate. Stimulation in the presence of MNI-glutamate significantly increased condensate signal at 20- (p = 0.0485, post hoc Tukey) and 30-min (p = 0.0214, post hoc Tukey) post-stimulation. **p < 0.01, paired t-test
Article Snippet: Organotypic slices were submerged in a low Mg 2+ HEPES buffer and images acquired at room temperature on a multiphoton system (Scientifica Hyperscope with a Coherent Chameleon Discovery; Nikon 16x, 0.8 NA lens or a Nikon 25x, 1.1 NA lens).
Techniques: Activity Assay, Expressing, Transfection, Imaging
