Journal: Nature structural & molecular biology

Article Title: Rational optimization of a transcription factor activation domain inhibitor.

doi: 10.1038/s41594-023-01159-5

Figure Lengend Snippet: Fig. 1 | AR phase separation is driven by tyrosine residues in the AD. a, Predicted structure of AR, colored by structure-prediction confidence from high (blue) to low (yellow). The domains and the native NLS are highlighted. b, Live-cell STED imaging of representative (n > 3) HEK293T cells transfected with AR constructs tagged with mEGFP. Cells were imaged after treatment with 10 nM DHT for 4 h. Scale bar, 5 μm. The dashed line indicates the nuclear periphery. c, Intensity of AR AD NMR resonances at different concentrations, relative to the intensity at 25 μM. The positions of Tau-1, Tau-5 and 23FQNLF27 are highlighted. Green circles indicate residues that were not visible (NV) or not assigned (NA), including residues in polyglutamine (pQ), polyproline (pP) and polyglycine (pG) tracts. Yellow and orange circles represent the positions of tyrosine residues substituted by serines in 8YtoS and 14YtoS; all tyrosine residues were substituted in 22YtoS. d, Fluorescence microscopy images of 40 µM AR-AD droplets (WT* and mutants) at 1 M NaCl. Scale bar, 10 μm. e, Scheme of the phase diagram

Article Snippet: HEK293T and HeLa cells were imaged on a Leica Stellaris STED DMI 8 microscope equipped with an okolab incubation chamber set to 37 °C and a constant concentration of CO2 (5%). eGFP imaging was performed using a 473-nm stimulation wavelength laser at 20% power and a 592-nm depletion laser at 20% power.

Techniques: Imaging, Transfection, Construct, Fluorescence, Microscopy