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

FIGURE 3.

FIGURE 3

Slingshot homolog-1 (SSH1)-mediated p62 inhibition at pSer403 increases small EV secretion. (A) Representative images of DiI-stained small EVs secreted from NIH3T3 cells transfected with control siRNA or SSH1 siRNA. (B) Quantification of secreted small EVs for (A). Data are presented as means ± SEM. n = 3 independent experiments, t-test, ***P = 0.0005. (C) Representative immunoblots for SSH1 and β-Actin protein levels in cell lysates for (A). (D) Representative images of DiI-stained small EVs secreted from NIH3T3 cells expressing VC or Flag-SSH1 (E) Quantification of secreted small EVs for (D). Data are presented as means ± SEM. n = 3 independent experiments, t-test, #P < 0.0001 (F) Representative immunoblots for Flag and β-Actin protein levels in cell lysates for (D). (G) Representative images of DiI-stained small EVs secreted from NIH3T3 cells co-transfected with VC or Flag-SSH1 and control siRNA or p62 siRNA. (H) Quantification of secreted small EVs for (G). Data are presented as means ± SEM. n = 3 independent experiments, 2-way ANOVA, followed by Sidak’s post-hoc, ***P = 0.0004. n.s. = not significant. (I) Representative immunoblots showing SSH1, p62, and β-Actin protein levels in cell lysates for (G). (J) Representative images of DiI-stained small EVs secreted from NIH3T3 cells co-transfected with VC or Flag-SSH1 and GFP, GFP-p62, or GFP-p62 S403E. (K) Quantification of secreted small EVs for (J). Data are presented as means ± SEM. n = 3 independent experiments, 2-way ANOVA, followed by Sidak’s post-hoc, ***P = 0.0007, **P = 0.001. n.s. = not significant. (L) Representative immunoblots for SSH1, p62, and β-Actin protein levels in cell lysates for (J).