Fig. 2: F-actin induces the recruitment of MICAL1 to the GPIb-IX-V complex upon platelet activation. | Nature Communications

Fig. 2: F-actin induces the recruitment of MICAL1 to the GPIb-IX-V complex upon platelet activation.

From: F-actin disassembly by the oxidoreductase MICAL1 promotes mechano-dependent VWF-GPIbα interaction in platelets

Fig. 2: F-actin induces the recruitment of MICAL1 to the GPIb-IX-V complex upon platelet activation.

a Identification of MICAL1 as a potential actin-depolymerizing protein interacting with GPIbα/GPIbβ thanks to the database BioGRID4.4 and the Gene Ontology “Actin filament depolymerization” (GO: 0030042). Created with BioRender.com. b Lysates of human platelets treated or not with LatA (15 µM) at 0 s (resting platelets), after 30 s or 2 min of activation (VWF 10 mg/mL/Risto 0.4 mg/mL) were separated into Triton X-100 soluble and insoluble fractions and blotted. Quantification of insoluble MICAL1 (mean ± SD, N = 4 independent experiments from 4 different donors, two-way ANOVA with Šídák post hoc test, F(2, 18) = 27.06, 2 min: p = 0.00000003), GPIbα (mean ± SD, N = 3 independent experiments from 4 different donors, two-way ANOVA, F(2, 12) = 49.40, 2 min: p = 0.00000005) and β-actin (mean ± SD, N = 4 independent experiments from 4 different donors, two-way ANOVA with Šídák post hoc test, F(2, 18) = 2.424, 2 min: p = 0.0000000004). Left panel: representative western blots. Right panels: MICAL1, GPIbα, β-actin quantification from western blots. c GPIbα immunoprecipitation from lysates from washed human platelets in resting or activated conditions (VWF 10 mg/mL/Risto 0.4 mg/mL) human platelets treated with LatA (15 µM) or DMSO as control and blotted for GPIbα, β-actin, (mean ± SD, N = 5 independent experiments from 4 different donors, one-way ANOVA with Šídák post hoc test, F = 202, df = 4) and MICAL1 (mean ± SD, N = 5 independent experiments from 4 different donors, one-way ANOVA with Šídák post hoc test, F = 19.2, df = 4). Left Panel: representative western blot. Right panel: β-actin and MICAL1 quantification from western blots. d Correlation of the presence of MICAL1 and β-actin associated to GPIbα after immunoprecipitation from experiments performed in (c). R2 and p were obtained through simple linear regression. Vertical dotted lines indicate a repositioned gel lane. Source data are provided as a Source Data file.

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