Skip to main content
. 2022 Aug 25;14:933979. doi: 10.3389/fnagi.2022.933979

FIGURE 4.

FIGURE 4

Slingshot homolog-1 (SSH1) increases small EV secretion partially through cofilin activation and F-actin disruption. (A) Schematic of SSH1 full length, SSH1ΔC, and SSH1ΔN proteins. (B) Representative images of DiI-stained small EVs secreted from NIH3T3 cells expressing VC, Flag-SSH1, Flag-SSH1ΔN, or CFP-SSH1ΔC. (C) Quantification of secreted small EVs for (B). Data are presented as means ± SEM. n = 3 independent experiments, 1-way ANOVA, followed by Dunnett’s post-hoc, #P < 0.0001, ***P = 0.001, **P = 0.0069. (D) Representative immunoblots for Flag, GFP, and β-Actin protein levels in cell lysates for (B). (E) Representative images of DiI-stained small EVs secreted from NIH3T3 cells expressing GFP or GFP-Cofilin. (F) Quantification of secreted small EVs for (E). Data are presented as means ± SEM. n = 3 independent experiments, t-test, #P < 0.0001. (G) Representative immunoblots for cofilin and β-Actin protein levels in cell lysates for (E). (H) Representative images of DiI-stained small EVs secreted from NIH3T3 cells expressing GFP, GFP-cofilin-S3A, or GFP-cofilin-S3E. (I) Quantification of secreted small EVs for (H). Data are presented as means ± SEM. n = 3 independent experiments, 1-way ANOVA, followed by Dunnett’s post-hoc, #P < 0.0001. (J) Representative immunoblots for GFP and β-Actin protein levels in cell lysates for (H). (K) Representative images of DiI-stained small EVs secreted from NIH3T3 cells transfected with control siRNA or cofilin siRNA. (L) Quantification of secreted small EVs for (K). Data are presented as means ± SEM. n = 3 independent experiments, t-test, #P < 0.0001. (M) Representative immunoblots for cofilin and β-Actin protein levels in cell lysates for (K). (N) Representative images of DiI-stained small EVs secreted from NIH3T3 cells with or without 100 nM Latrunculin B (LatB) or 100 nM Jasplakinolide (Jasp) treatment for 8 h. (O) Quantification of secreted small EVs for (N). Data are presented as means ± SEM. n = 3 independent experiments, 1-way ANOVA, followed by Dunnett’s post-hoc, #P < 0.0001. n.s. = not significant.