Figure 7.

Preventing GlcNAc from Incorporating into the N-Glycan Branching Pathway Blocks GlcNAc Effects on Fate Potential

(A) Experimental design: Undifferentiated E12 NSPCs were treated with either 0.5 μM Kif for 4 days, 80 mM GlcNAc for 3 days, or pre-treated with Kif 1 day prior to an additional 3 days of Kif + GlcNAc supplementation (Combo). Cells were differentiated for 3 days after treatment.

(B) L-PHA flow cytometry analysis of cells after the 4 days of treatment indicates significantly lower highly branched N-glycans on Kif- and Combo-treated cells compared with untreated controls (untreated versus Kif or Combo p < 0.0001, one-way ANOVA, Tukey post hoc for multiple comparisons).

(C) No difference in neuronal differentiation between Kif- and Combo-treated cells, indicating no effect of GlcNAc in the presence of Kif. Combo-treated cells show a slight decrease in neuron formation compared with untreated controls (untreated versus Combo p = 0.0311, one-way ANOVA, Tukey post hoc for multiple comparisons).

(D) When blocked by Kif, GlcNAc has no effect on astrocyte formation since no difference was observed between Kif- and Combo-treated samples.

Error bars SEM. N = 3 or more independent biological repeats (^∗p < 0.05, ^∗∗∗∗p < 0.0001).