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. Author manuscript; available in PMC: 2021 Aug 13.
Published in final edited form as: Nat Metab. 2021 May 24;3(5):714–727. doi: 10.1038/s42255-021-00390-y

Figure 5: RhoA induces glucose transporter SLC2A3 translocation which drives subcellular glycolysis in thrombin stimulated contractile endothelial cells.

Figure 5:

a, Western blot of phospho-cofilin and phospho-MLC in the presence or absence of siRNA targeted towards SLC2A3 and thrombin treatment (n = 4 all; biological replicates, p = 0.0140 for pCFL and p = 0.0233 for ppMLC). Loading control (ACTB) for each protein denoted in parentheticals. Original blot in Source Data b. b, LPR of siRNA targeted towards SLC2A3 or control with and without thrombin treatment (n = 277, 263, 275, 278 from left to right, p < 0.0001 for all comparisons). c, Representative immunofluorescence of membrane markers CDH5 and SLC2A3 in thrombin treated cells causes SLC2A3 granule localization to membrane spikes, which is inhibited by Y27632, by TIRF microscopy. Scale bar = 5 μm. d, Quantification of SLC2A3 granule brightness near the membrane of (c) (a.u. = arbitrary units, n = 36, 36, 17 images from left to right. p = 0.003 for thrombin vs. control, and p = 0.0499 for thrombin vs thrombin + Y27632). e, TEER measurement of thrombin-induced permeability changes in the presence or absence of siRNA targeted towards SLC2A3 (n = 4; biological replicates, p = 0.0248). f, Representative immunofluorescence of CDH5 and intercellular gaps, as quantified by gap size index, in HAECs treated with control or siRNA targeted towards SLC2A3 followed by thrombin treatment (n = 12, 16, 8, 32 images from left to right, p = 0.0184). Scale bar = 150 μm. Statistical significance determined by multiple unpaired two-tailed t-test with multiple comparisons adjustment by the Holm-Šídák method (a), two-sided Welch’s t-test (e, f) or by one-way ANOVA followed by Bonferroni test (b, d). All error bars are SEM.