Figure 6.
SNX17 and SNX27 define two distinct pathways for structural plasticity by recycling specific cargoes. (A) Rat hippocampal neurons were co-transfected at DIV12 with either ctrl- or SNX17-shRNA and the indicated constructs. At DIV16, neurons were either treated with cLTP or left untreated and fixed 50 min after cLTP. The width of dendritic spines in the first 30 µm of secondary dendrites were quantified. ctrl-shRNA + eGFP: 0.393 ± 0.011, N = 30 neurons; ctrl-shRNA + eGFP with cLTP: 0.515 ± 0.012, N = 30 neurons; SNX17-shRNA + eGFP: 0.370 ± 0.006, N = 30 neurons; SNX17-shRNA + eGFP with cLTP: 0.334 ± 0.014, N = 30 neurons; SNX17-shRNA + SNX17-R: 0.373 ± 0.008, N = 30 neurons; SNX17-shRNA + SNX17-R with cLTP: 0.509 ± 0.012, N = 30 neurons; SNX17-shRNA + SNX27-R: 0.367 ± 0.006, N = 30 neurons; SNX17-shRNA + SNX27-R with cLTP: 0.356 ± 0.006, N = 30 neurons. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (B) Rat hippocampal neurons were co-transfected at DIV12 with either ctrl- or SNX27-shRNA and the indicated constructs. At DIV16, neurons were either treated with cLTP or left untreated and fixed 50 min after cLTP. The width of dendritic spines in the first 30 µm of secondary dendrites were quantified. ctrl-shRNA + eGFP: 0.409 ± 0.008, N = 30 neurons; ctrl-shRNA + eGFP with cLTP: 0.503 ± 0.013, N = 30 neurons; SNX27-shRNA + eGFP: 0.402 ± 0.008, N = 30 neurons; SNX27-shRNA + eGFP with cLTP: 0.398 ± 0.010, N = 30 neurons; SNX27-shRNA + SNX27-R: 0.397 ± 0.009, N = 30 neurons; SNX27-shRNA + SNX27-R with cLTP: 0.478 ± 0.010, N = 30 neurons; SNX27-shRNA + SNX17-R: 0.388 ± 0.009, N = 30 neurons; SNX27-shRNA + SNX17-R with cLTP: 0.380 ± 0.006, N = 30 neurons. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (C) DIV11 rat cortical neurons were infected with lentiviruses carrying ctrl-, SNX17-, or SNX27-shRNAs, and the surface levels of β1-integrin and GluA1 were determined at DIV17 using surface biotinylation assays. Knockdown of SNX17 and SNX27 was validated by western blot of the lysate, and GAPDH was used as a loading control. (D) The levels of surface β1-integrin protein were quantified and expressed as a percentage of ctrl-shRNA. ctrl-shRNA: 100%, SNX17-shRNA: 48.350 ± 4.901%, SNX27-shRNA: 129.400 ± 6.202%. N = 4 independent experiments. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (E) The levels of surface GluA1 protein were quantified and expressed as a percentage of ctrl-shRNA. ctrl-shRNA: 100%, SNX17-shRNA: 113.200 ± 5.011%, SNX27-shRNA: 51.820 ± 0.696%. N = 3 independent experiments. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. DIV, days in vitro. Source data are available for this figure: SourceData F6.

SNX17 and SNX27 define two distinct pathways for structural plasticity by recycling specific cargoes. (A) Rat hippocampal neurons were co-transfected at DIV12 with either ctrl- or SNX17-shRNA and the indicated constructs. At DIV16, neurons were either treated with cLTP or left untreated and fixed 50 min after cLTP. The width of dendritic spines in the first 30 µm of secondary dendrites were quantified. ctrl-shRNA + eGFP: 0.393 ± 0.011, N = 30 neurons; ctrl-shRNA + eGFP with cLTP: 0.515 ± 0.012, N = 30 neurons; SNX17-shRNA + eGFP: 0.370 ± 0.006, N = 30 neurons; SNX17-shRNA + eGFP with cLTP: 0.334 ± 0.014, N = 30 neurons; SNX17-shRNA + SNX17-R: 0.373 ± 0.008, N = 30 neurons; SNX17-shRNA + SNX17-R with cLTP: 0.509 ± 0.012, N = 30 neurons; SNX17-shRNA + SNX27-R: 0.367 ± 0.006, N = 30 neurons; SNX17-shRNA + SNX27-R with cLTP: 0.356 ± 0.006, N = 30 neurons. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (B) Rat hippocampal neurons were co-transfected at DIV12 with either ctrl- or SNX27-shRNA and the indicated constructs. At DIV16, neurons were either treated with cLTP or left untreated and fixed 50 min after cLTP. The width of dendritic spines in the first 30 µm of secondary dendrites were quantified. ctrl-shRNA + eGFP: 0.409 ± 0.008, N = 30 neurons; ctrl-shRNA + eGFP with cLTP: 0.503 ± 0.013, N = 30 neurons; SNX27-shRNA + eGFP: 0.402 ± 0.008, N = 30 neurons; SNX27-shRNA + eGFP with cLTP: 0.398 ± 0.010, N = 30 neurons; SNX27-shRNA + SNX27-R: 0.397 ± 0.009, N = 30 neurons; SNX27-shRNA + SNX27-R with cLTP: 0.478 ± 0.010, N = 30 neurons; SNX27-shRNA + SNX17-R: 0.388 ± 0.009, N = 30 neurons; SNX27-shRNA + SNX17-R with cLTP: 0.380 ± 0.006, N = 30 neurons. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (C) DIV11 rat cortical neurons were infected with lentiviruses carrying ctrl-, SNX17-, or SNX27-shRNAs, and the surface levels of β1-integrin and GluA1 were determined at DIV17 using surface biotinylation assays. Knockdown of SNX17 and SNX27 was validated by western blot of the lysate, and GAPDH was used as a loading control. (D) The levels of surface β1-integrin protein were quantified and expressed as a percentage of ctrl-shRNA. ctrl-shRNA: 100%, SNX17-shRNA: 48.350 ± 4.901%, SNX27-shRNA: 129.400 ± 6.202%. N = 4 independent experiments. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. (E) The levels of surface GluA1 protein were quantified and expressed as a percentage of ctrl-shRNA. ctrl-shRNA: 100%, SNX17-shRNA: 113.200 ± 5.011%, SNX27-shRNA: 51.820 ± 0.696%. N = 3 independent experiments. Statistical significance was determined using one-way ANOVA with Tukey’s post hoc test, ****P < 0.001. Error bars are SEM. DIV, days in vitro. Source data are available for this figure: SourceData F6.

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