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. 2021 Apr 15;10:e61401. doi: 10.7554/eLife.61401

Figure 1. Plin4 AH forms very stable oil particles.

(A) Helical wheel representations of Plin4 and Plin4 4T > S AHs. (B) Summary of LD localization of Plin4 AH and mutants (Čopič et al., 2018). (C) A 10 µl drop of triolein was added to 190 μl of HK buffer containing the indicated proteins (Plin4 12mer, Plin4 4mer or 4T > S, all at 0.5 mg/ml). After vigorous vortexing, the samples were photographed. (D) Dynamics of Plin4 12mer interaction with oil as assessed by FRAP assays. Emulsions of triolein with unlabeled Plin4 12mer (0.5 mg/ml) and Alexa488-labeled Plin4 12mer (0.025 mg/ml) 12mer were prepared as in C and visualized by fluorescence microscopy. FRAP was performed on large droplets, which were entirely bleached (top row), or in the bulk as a control (middle row). The lower row shows a FRAP experiment performed on a limited region of the droplet. A summary of all FRAP experiments is shown in Figure 1—figure supplement 1B. (E) FRAP of Plin4 4mer (0.5 mg/ml) mixed with Alexa488-labeled Plin4 4mer (0.01 mg/ml). (F) Light microscopy images of Plin4 12mer and 4T > S emulsions at different time points after preparation by vortexing. (G) Size distribution as assessed by dynamic light scattering (DLS) of a Plin4 12mer/triolein emulsion from 3 hr to 28 days after the vortexing reaction. Experiment was repeated two times. Scale bars: 5 µm.

Figure 1.

Figure 1—figure supplement 1. Summary of FRAP experiments on oil droplets.

Figure 1—figure supplement 1.

(A) Examples of FRAP experiments on oil droplets formed with Plin4 4T > S mutant. Emulsions of triolein were prepared with a solution of 4T > S (0.5 mg/ml) and Alexa488-4T > S (0.025 mg/ml). FRAP was performed using the same protocol as for Plin4 12mer. Note that there were very few droplets present in this suspension. Scale bar: 5 µm. (B) Summary of the results of all FRAP experiments on oil droplets formed with Plin4 12mer, Plin4 4mer or 4T > S (4mer) mutant.