Figure 4. Effects of BuGZ phase separation on AurA binding. (A and B) Purified YFP-xBuGZ or YFP-xBuGZΔN58 was incubated to allow coacervation in the presence of purified mCherry-xAurA (A) or mCherry (B). Bars, 10 µm. (C) Purified YFP, YFP-xBuGZ, or YFP-xBuGZ13S were incubated with purified GST or GST-xAurA at RT or 4°C followed by GST pulldown, Western blotting (0.1% of total reaction and 10% of total YFP-xBuGZ and YFP-xBuGZ13S pulldowns were analyzed), and Coomassie Brilliant blue (CBB) staining (1% of the total reaction and 10% of total pulldowns were analyzed). (D) Schematic of droplet spindown assay. (E–G) Quantification of YFP-xBuGZ and xAurA (E), YFP-xBuGZΔN58 and xAurA (F), and YFP-xBuGZ13S and xAurA (G) in the pellets (X axis) against the initial input xBuGZ proteins (Y axis). Error bars indicate SEM.
Figure 4.

Effects of BuGZ phase separation on AurA binding. (A and B) Purified YFP-xBuGZ or YFP-xBuGZΔN58 was incubated to allow coacervation in the presence of purified mCherry-xAurA (A) or mCherry (B). Bars, 10 µm. (C) Purified YFP, YFP-xBuGZ, or YFP-xBuGZ13S were incubated with purified GST or GST-xAurA at RT or 4°C followed by GST pulldown, Western blotting (0.1% of total reaction and 10% of total YFP-xBuGZ and YFP-xBuGZ13S pulldowns were analyzed), and Coomassie Brilliant blue (CBB) staining (1% of the total reaction and 10% of total pulldowns were analyzed). (D) Schematic of droplet spindown assay. (E–G) Quantification of YFP-xBuGZ and xAurA (E), YFP-xBuGZΔN58 and xAurA (F), and YFP-xBuGZ13S and xAurA (G) in the pellets (X axis) against the initial input xBuGZ proteins (Y axis). Error bars indicate SEM.

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