Figure 6

Overexpression of DNMT3a alters GRN promoter activity in primary cortical neurons and reduces GRN mRNA expression in LCLs. (a) pCpGL plasmid containing the GRN core promoter and a DNMT3a overexpression construct were transiently transfected in HEK 293FT cells (left panel) and in rat primary cortical neurons (right panel). Relative luciferase activity was determined by normalizing firefly luciferase against Renilla luciferase activity. Empty vectors were used as negative control. Firefly luciferase expression was significantly reduced upon DNMT3a overexpression. Mean ± SEM, n ≥ 3. ***p < 0.001, ANOVA with Tukey’s Multiple Comparison test. (b) Lentiviral expression of DNMT3a in LCLs #3 and #14. Overexpression was verified by qPCR five days after viral transduction. n = 5, mean ± SEM, *p < 0.05, Student’s t-test. (c) GRN mRNA expression levels were significantly reduced in DNMT3a overexpressing LCLs as quantified by qPCR and normalized to PGK1 expression levels. n = 5, mean ± SEM, *p < 0.05, Student’s t-test.