Extended Data Fig. 3: MR dynamics in living cells show propensity for LLPS and condensate formation that is driven by the intrinsic disordered protein region.
From: Mineralocorticoid receptor phase separation modulates cardiac preservation
a, Neonatal rat cardiomyocytes (NRCM) expressing green fluorescent protein (GFP)-MR using adenovirus were bathed in cold (4°C) HTK preservation solution under hypoxic conditions (1% O2) for 10 h. A laser was used to perform fluorescence recovery after photobleaching (FRAP) within the nucleus. The dark region in the middle represents the nucleoli. Photobleached regions are circled and fluorescence recovery was quantified in adjoining graph. Data are presented as means ± SD (n = 3 biological replicates). b, We designed constructs that used lipofectamine to transfect Human Embryonic Kidney (HEK-293) cell cultures to express a mCherry-optoDrop (OptoDroplets) vector with either a full length MR (MR_FL) protein or a mutant MR without the IDR at the N-terminal domain (MRΔIDR). c. The HEK-293 cells were then immersed in histidine-tryptophan-ketoglutarate preservation solution under normoxic conditions and exposed to blue light for up to 60 s to induce MR condensate formation (white arrow). MR_FL is seen throughout the entire cell whereas MRΔIDR remained mostly within the cytoplasm thus outlining the nucleus. Data shown represent n = 4 biological replicates. d, Laser “photobleaching” of MR_FL condensates in live HEK-293 cells followed by fluorescence recovery. Photobleached region is circled. Image shown represent n = 4 biological replicates.
