XB-IMG-170413
Xenbase Image ID: 170413
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Figure 7. Focal Hotspots of Nascent β-actin at Axonal Branch Points and within Branches In Vivo(A) Fluorescence recovery after photobleaching experiment of Venus constructs in vivo.(B) Fluorescence heatmaps illustrating a ubiquitous recovery pattern for the Venus control construct. In contrast, Venus-β-actin signal recovered in hotspots at branch points and within branches. The formation of these hotspots was inhibited by cycloheximide (CHX), indicating de novo synthesis and accumulation of β-actin in highly specific focal points in RGC axons.(C) An example of multiple Venus-β-actin hotspots forming at different sub-compartments of a branch. Kymograph displays the 300 s FRAP from the branch tip to axon shaft as indicated by the magenta arrow. At least four distinct hotspots can be identified in this single branch.(D) The fluorescence variation index (FVI) is defined by normalizing the standard deviation (SD) of fluorescence in branches (Fbranch) to the SD of fluorescence in axon shaft (Fshaft).(E) FVI after FRAP over the course of 5 min. Dotted lines represent least-squares fits to a quadratic function. (Venus control versus Venus-β-actin: F3,6594 = 396, p < 0.0001; Venus-β-actin versus Venus-β-actin + CHX: F3,5994 = 466.7, p < 0.0001). Inset displays the differences between the conditions. Error bars represent SEM. ∗∗∗p < 0.001, ###p < 0.001 (extra sum-of-squares F test for E). Scale bars, 5 μm. Image published in: Wong HH et al. (2017) © 2017 The Authors. This image is reproduced with permission of the journal and the copyright holder. This is an open-access article distributed under the terms of the Creative Commons Attribution license Larger Image Printer Friendly View |