Extended Data Fig. 3: Hexanediol treatment selectively disrupts C. albicans TF condensates even during co-compartmentalization with other TFs.

a, Images of Efg1, Czf1, Wor1 (CaCmWor1), and Wor4 droplets at the indicated concentrations with or without 10% 1,6- or 2,5-hexanediol. For hexanediol treatment, proteins were incubated with TEV for 30 minutes in 10 mM Tris-HCl, pH 7.4, 150 mM NaCl, at 22 °C, and then mixed with 1,6- or 2,5-hexanediol in the same buffer, incubated for 10 minutes, and imaged. Wor1, Wor4, and Czf1 assays also included 5% PEG-8000. Where indicated for Wor4, hexanediol was added for 10 minutes and then TEV/PEG-8000 added and the sample incubated for an additional 30 minutes prior to imaging. Images represent a single experimental replicate with assays repeated at least twice with similar results. Scale bars; 10 μm. b, Representative images of fluorescently labeled Efg1, Wor1 (CaWor1), Wor4, and Czf1 proteins compartmentalized within Efg1 condensates, and treated with 10% 1,6- or 2,5-hexanediol. Unlabeled bulk protein (15 μM) was mixed with each of the fluorescently labeled proteins (37.5 nM) in 10 mM Tris-HCl, pH 7.4, 150 mM NaCl. Proteins were then incubated at 22 °C with TEV for 30 minutes and treated with 1,6- or 2,5-hexanediol in the same buffer for 10 minutes prior to imaging. Dylight NHS-Ester labeling of the 4 proteins used fluors of 405, 488, 550 and 633 nm. Images represent a single experimental replicate with assays performed three times with similar results. Scale bar, 10 μm; images are maximum Z-stack projections. c, Representative images of fluorescently labeled Efg1, Wor1 (CaWor1), Wor4, and Czf1 proteins compartmentalized within Czf1, Wor1(CaCmWor1), or Wor4 condensates. Unlabeled bulk proteins (15 μM) were mixed with each of the fluorescently labeled proteins (37.5 nM) in 10 mM Tris-HCl, pH 7.4, 150 mM NaCl. Proteins were then incubated at 22 °C with TEV for 30 min. Dylight NHS-Ester labeling of the 4 proteins used fluors of 488, 550, 405, and 633 nm. Images represent a single experimental replicate, with assays performed three times with similar results. Scale bars, 10 μm; images are maximum Z-stack projections.