Fig. 2: TPM4 can undergo phase separation.
From: TPM4 condensates glycolytic enzymes and facilitates actin reorganization under hyperosmotic stress
a Left: representative images of MDA-MB-231 cells expressing low, medium and high levels of TPM4-AcGFP. Scale bars, 20 μm. Top right: IDR (red) and non-IDR (blue) regions of TPM4 according to MobiDB (https://mobidb.bio.unipd.it/); bottom right: residue-based PDP of TPM4 according to FuzDrop (https://fuzdrop.bio.unipd.it/predictor). b Representative images of purified TPM4-AcGFP protein in storage and phase separation buffer. Scale bars, 10 μm. Zoomed images showing the fusion of TPM4 droplets in vitro. c Representative images of TPM4-AcGFP droplets with different protein concentrations (0.5 μM, 10 μM, 50 μM) in phase separation buffer. Scale bar, 10 μm. d Left: representative images of MDA-MB-231 cells expressing TPM4-mCherry-Cry2 upon blue light exposure and withdrawal. Scale bar, 20 μm. Right: zoomed images showing the formation and fusion of TPM4 condensates upon blue light exposure within 2 min. e Left: representative images of MDA-MB-231 cells expressing TPM4-AcGFP under 100 mM sorbitol treatment and washout. Scale bar, 20 μm. Right: zoomed images showing the fusion events of TPM4 condensates. f Representative time-lapse images of MDA-MB-231 cells expressing TPM4-AcGFP treated with 12.5 mM, 25 mM, 50 mM sorbitol within 2 min. Scale bar, 20 μm. g Representative time-lapse images of TPM4-AcGFP and LifeAct-mScarlet in MDA-MB-231 cells treated with 100 mM sorbitol within 20 s. Scale bar, 2 μm. h Schematic illustration of CRISPR-Cas9-mediated endogenous tagging of TPM4 gene with GFP at C-terminus. i Representative images of endogenous TPM4-AcGFP in MDA-MB-231 cells before (isosmotic) and after (hyperosmotic) 100 mM sorbitol treatment for 3 min. Scale bar, 20 μm. j Representative images of endogenous TPM4-AcGFP in MDA-MB-231 cells before and after 2.5 μM latrunculin B treatment for 1 h. Scale bar, 20 μm.
