Journal: Virologica Sinica

Article Title: PABP-driven secondary condensed phase within RSV inclusion bodies activates viral mRNAs for ribosomal recruitment

doi: 10.1016/j.virs.2023.12.001

Figure Lengend Snippet: The viral mRNP granules within RSV IBs are secondary biomolecular condensates formed by LLPS. A – C Live imaging of HeLa-GFP-PABP cells infected with RSV. The enlargement of white-framed regions was shown on the right. The time point is indicated at the top right corner of each frame ( A and B ). A The dynamic behavior of viral mRNP granules. Arrowhead indicates a cluster of viral mRNP granules released and diluted into the cytoplasm in a to f order, also referred to Video S1. Scale bar, 5 ​μm. B Representative images showing the fusion of two touching viral mRNP granules (a and b) labeled with GFP-PABP into larger spherical granules. Also referred to as Video S2. Scale bar, 2 ​μm. C Fluorescence recovery after photobleaching (FRAP) analysis of viral mRNP granules labeled with GFP-PABP. The bleached region of interest (ROI) is marked with a white circle. The FRAP data from four distinct granules in four distinct cells were fitted into a single curve (blue) by using R/Rstudio. Also referred to as Video S3. Scale bar, 2 ​μm. D HeLa-GFP-PABP cells were treated with 1.5% 1,6-hexanediol for 5 ​min after being infected with RSV for 36 ​h. IBs were detected by mouse anti-RSV P antibody (red). Arrowhead indicates viral mRNP granules dissolved by 1,6-hexanediol. Nuclei are stained with DAPI (blue in Merge). Scale bar, 10 ​μm.

Article Snippet: The FRAP data from four distinct granules in four distinct cells were fitted into a single curve (blue) by using R/Rstudio.

Techniques: Imaging, Infection, Labeling, Fluorescence, Staining

Journal:

Article Title: The polarity and dynamics of microtubule assembly in the budding yeast Saccharomyces cerevisiae

doi: 10.1038/71357

Figure Lengend Snippet: Statistical analysis of FSM and FRAP marks relative to SPBs

Article Snippet: Statistical analysis of FSM and FRAP data Metamorph software was used to measure selected parameters (distance or fluorescence intensity) from time series that were suitable for analysis.

Techniques:

Journal:

Article Title: The polarity and dynamics of microtubule assembly in the budding yeast Saccharomyces cerevisiae

doi: 10.1038/71357

Figure Lengend Snippet: a, A metaphase spindle in a cell expressing GFP–tubulin that was photobleached in the lower half-spindle and then observed by time-lapse microscopy. The −7 s time point shows the spindle before photobleaching. The laser was targeted to the lower half of the spindle (white arrowhead), which was exposed to the laser for 25 ms. Time point 0 shows that the upper half-spindle is still fluorescent (grey arrowhead) while the lower half has been bleached. Time-lapse analysis for 20 min revealed that the bleached portion of the spindle recovered fluorescence (white arrowhead), while the unbleached portion lost intensity, because of photobleaching by fluorescent exposures for image acquisition and incorporation of bleached tubulin subunits. Time points are given in seconds. b, The graph shows integrated intensity measurements of a 5 pixel × 5 pixel square placed over the bleached region (squares) and the unbleached region (diamonds) over time. These values are corrected for photobleaching that occurred during time-lapse image acquisition; a decrease from the initial value of ~20% had occurred by the 360-s time point. Also plotted are the predicted rates of FRAP for bleached (grey line) and unbleached (black line) half-spindles. These theoretical values were derived using the first-order rate constant k, calculated as described in Methods. Scale bar in a represents 2 μm.

Article Snippet: Statistical analysis of FSM and FRAP data Metamorph software was used to measure selected parameters (distance or fluorescence intensity) from time series that were suitable for analysis.

Techniques: Expressing, Time-lapse Microscopy, Fluorescence, Derivative Assay

Journal: Cerebral cortex (New York, N.Y. : 1991)

Article Title: Homeostatic regulation of gephyrin scaffolds and synaptic strength at mature hippocampal GABAergic postsynapses.

doi: 10.1093/cercor/bhs260

Figure Lengend Snippet: Figure 4. FRAP rates of GFP-gephyrin clusters are slowed during inhibitory synapse maturation in vitro. (A) Series of FRAP images taken from single GFP-gephyrin clusters in the CA1 stratum radiatum after bleaching at the 0-min time point; images are from 1- and 4-weeks-old slice cultures prepared from Thy1-EGFPgephyrin mice. Control, unbleached GFP-gephyrin cluster. (B) Quantitative evaluation of experiments as shown in (A). Fast FRAP was seen in 1-week-old cultures, with 30% of the averaged prebleach GFP-gephyrin fluorescence recovering within the first 5 min after photobleaching. In contrast, in 4-weeks-old cultures, 30% recovery was reached only 30 min after bleaching (n = 5 cultures per stage; 5–12 clusters bleached per culture; significance indicated for averaged 65–80 min values). (C) Group parameters obtained by bi-exponential fitting of the FRAP data shown in (B). While the τ values of the slow and the fast FRAP components were not significantly different (n.s.) between 1- and 4-weeks-old cultures, the relative fraction of the fast component was lower, and of the slow component larger, at 4 weeks when compared with 1 week. In addition, a significant increase in the stationary fraction of gephyrin clusters was observed in

Article Snippet: FRAP data were fitted using the curve fitting toolbox of MATLAB version 7.5 (The MathWorks Inc., Natick, MA, United States of America) with the following bi-exponential equation: f ðtÞ 1⁄4 Pfð1 e t=tf Þ þ Psð1 e t=ts Þ where Pf denotes the fraction of the fast component with the recovery time constant τf and Ps the fraction of the slow component with the recovery time τs, respectively.

Techniques: In Vitro, Control