Additional characterization of G3BP1 lysate granules. (A) Cell pellets were resuspended in lysis buffer with or without 0.5% NP-40, and Hoechst was added. Quantification of Hoechst staining shows that a substantial portion of nuclei were lysed, and relatively few nuclei remain intact in the lysis buffer. Graph represents mean ± SD, n = 5 independent representative fields. (B) Western blot of U2OS or G3BP1-GFP U2OS lysates against purified G3BP1 standards. Cellular concentrations of G3BP1 were assessed by densitometry to be ∼100 nM endogenous G3BP1 and 150 nM G3BP1-GFP in G3BP1-GFP U2OS lysate, for a total of 250 nM total G3BP1 in G3BP1-GFP U2OS cells. (C) Lysate granules were treated with 10% 1,6-hexanediol at 15 or 30 min following induction by increasing concentration of G3BP1 by 20 µM using purified G3BP1, leading to clearance of the lysate granules over time. Granules were visualized by imaging G3BP1-GFP or DIC. Scale bar, 5 µm.
Figure S1.

Additional characterization of G3BP1 lysate granules. (A) Cell pellets were resuspended in lysis buffer with or without 0.5% NP-40, and Hoechst was added. Quantification of Hoechst staining shows that a substantial portion of nuclei were lysed, and relatively few nuclei remain intact in the lysis buffer. Graph represents mean ± SD, n = 5 independent representative fields. (B) Western blot of U2OS or G3BP1-GFP U2OS lysates against purified G3BP1 standards. Cellular concentrations of G3BP1 were assessed by densitometry to be ∼100 nM endogenous G3BP1 and 150 nM G3BP1-GFP in G3BP1-GFP U2OS lysate, for a total of 250 nM total G3BP1 in G3BP1-GFP U2OS cells. (C) Lysate granules were treated with 10% 1,6-hexanediol at 15 or 30 min following induction by increasing concentration of G3BP1 by 20 µM using purified G3BP1, leading to clearance of the lysate granules over time. Granules were visualized by imaging G3BP1-GFP or DIC. Scale bar, 5 µm.

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