Figure 3.
GJA1-20k inhibits actin polymerization. (A) Representative live-cell confocal imaging of actin, visualized by LifeAct-mCherry, in a HeLa cell expressing 0.5 µg of CP plasmid; scale bars = 10 µm. (B) Quantification of actin puncta density for HeLa cells transfected with GST-GFP, CP, or GJA1-20k-GFP. Data are presented as mean ± SD (n = 50–100 cells per group from three independent experiments). P > 0.05, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with multiple comparisons with Bonferroni’s post hoc test. (C) Representative TIRFM images (t = 15 min after initiation of polymerization) of cell-free Alexa Fluor 488–labeled actin in the presence of increasing concentrations of purified CP (alpha and beta subunits); scale bar = 5 µm. Reduction in actin polymerization is quantified by total fluorescence (A.U.) (D) Data are presented as mean ± SEM (n = 3 experiments per concentration). **P < 0.01, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with comparison to the actin only group with Bonferroni’s post hoc test. (E) Representative TIRFM images (t = 15 min after initiation of polymerization) of cell-free Alexa Fluor 488–labeled actin in the presence of increasing concentrations of GJA1-20k; scale bars = 5 µm. Reduction in actin polymerization is quantified by total fluorescence (A.U.) (F) Data are presented as mean ± SEM (n = 4–5 independent experiments per concentration) **P < 0.01, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with comparison to the actin only group with Bonferroni’s post hoc test.

GJA1-20k inhibits actin polymerization. (A) Representative live-cell confocal imaging of actin, visualized by LifeAct-mCherry, in a HeLa cell expressing 0.5 µg of CP plasmid; scale bars = 10 µm. (B) Quantification of actin puncta density for HeLa cells transfected with GST-GFP, CP, or GJA1-20k-GFP. Data are presented as mean ± SD (n = 50–100 cells per group from three independent experiments). P > 0.05, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with multiple comparisons with Bonferroni’s post hoc test. (C) Representative TIRFM images (t = 15 min after initiation of polymerization) of cell-free Alexa Fluor 488–labeled actin in the presence of increasing concentrations of purified CP (alpha and beta subunits); scale bar = 5 µm. Reduction in actin polymerization is quantified by total fluorescence (A.U.) (D) Data are presented as mean ± SEM (n = 3 experiments per concentration). **P < 0.01, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with comparison to the actin only group with Bonferroni’s post hoc test. (E) Representative TIRFM images (t = 15 min after initiation of polymerization) of cell-free Alexa Fluor 488–labeled actin in the presence of increasing concentrations of GJA1-20k; scale bars = 5 µm. Reduction in actin polymerization is quantified by total fluorescence (A.U.) (F) Data are presented as mean ± SEM (n = 4–5 independent experiments per concentration) **P < 0.01, ***P < 0.001, ****P < 0.0001 by one-way ANOVA with comparison to the actin only group with Bonferroni’s post hoc test.

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