Functional Role of SIL1 in Neurodevelopment and Learning

Figure 3

Expression of GluN2A was diminished after SIL1 silencing. (a) Two silencing vectors (shRNA#1 and shRNA#2) targeting SIL1 were designed and transfected into cultured cortical neurons at DIV6 through lentivirus transfection for two-day incubation, and whole cell lysates were harvested at DIV14. The scramble shRNA was used as the control. The expression of SIL1, GluN2A, GluN2B, and Bip was examined by Western blot. (b) Statistical analysis of (a): the expression of each protein was first normalized to GAPDH and then compared with the control. The levels of SIL1 and GluN2A were significantly reduced after shRNA#1 and shRNA#2 were reduced compared to the scramble shRNA-treated control group (SIL1: after shRNA#1 treatment, after shRNA#2 treatment; GluN2A: after shRNA#1 treatment, after shRNA#2 treatment). The levels of GluN2B and Bip were not altered (GluN2B: after shRNA#1 treatment, after shRNA#2 treatment; Bip: after shRNA#1 treatment, after shRNA#2 treatment). (c) The membrane expression of SIL1, GluN2A, GluN2B, and Bip was measured by biotinylation followed by Western blot (surface: membrane fraction; offer: total protein). (d) Statistical analysis of (c): the expression of each protein was first normalized to GAPDH and then compared with the control (GluN2A: after shRNA#1 treatment, after shRNA#2 treatment; GluN2B: after shRNA#1 treatment, after shRNA#2 treatment). All data were presented as . ^##. for all treatments.

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