Fig. 4: Solidifying mRNA-containing condensates strengthens translation repression.

A (Left) Schematic of the effect of different material properties on condensate function. (Right) Condensate composition is altered to accommodate optoFUS, a solid-like gel-forming light-activatable component. Inset: an image of the gel-like optoFUS cluster in a U2OS cell. B Example confocal images of the light-activated U2OS cell expressing the tagBFP reporter mRNA, optoMCP-FUS, and optoFUS. C Phase diagram of blue-light activated U2OS cells expressing optoMCP-FUS and optoFUS. Solid circles indicate cells with condensates while empty circles are those without condensates. D (Left) For individual cells, measured condensate compositions were visualized as circles of different radii. (Right) The measured composition of condensates in cells with different levels of optoMCP-FUS and optoFUS. A region with a dashed boundary near the origin corresponds to the concentration range where no phase separation was observed as in (C). E (Left) Representative confocal images of cells from each population in (D). Cell boundaries are indicated with yellow curves. (Right) Intensity profiles of optoMCP-FUS and optoFUS along white dashed lines. Fluorescence intensities were normalized with the maximum pixel values of optoMCP-FUS in each condensate. F (Left) Scatter plot for the normalized tagBFP intensities of individual cells after 12 h of blue light activation. A group of cells with strong translation repression, labeled as group 1, was chosen for further analysis. Cells in group 2, expressing only optoFUS, were chosen to have similar levels of clustering as those in group 1 (Supplementary Fig. 6F). (Right) 3D contour graph of the moving-averaged scatter plot. G (Top) Fluorescence images of condensates in two different groups in (F) during FRAP experiments. (Bottom) Fluorescence recovery curves for condensates in each group. Data are mean (solid line) ± SD (shaded area). n = 12 (Group 1) and 11 (Group 2). H Normalized tagBFP intensity after 12 h of light activation for cells in group 1 and 2. Data are mean ± SD. n = 18 (Group 1) and 12 (Group 2). *P < 0.05. P = 0.0102. Scale bars, 10 μm (A and B), 5 μm (E), and 1 μm (G). a.u., arbitrary units. Source data for panels (C, D, and F–H) are provided in the Source Data file.