Figure 1.

CRISPRa and lentivirus-mediated KCNJ6 expression similarly increase GIRK2 protein in human glutamatergic neurons. A, RT-qPCR test of 6 KCNJ6 gRNAs normalized to a scramble gRNA control in D42 NGN2 neurons (N = 1 donor, 2 replicates). B, RT-qPCR of KCNJ6 expression in D42 NGN2 neurons with scramble gRNA, no virus control, combined KCNJ6 gRNAs 1, 3, and 6 (CRISPRa) and lentiviral expression (Lenti; N = 1 donor, 3 replicates). C, Western blot of iso-CTL, CRISPRa, and Lenti expression of GIRK2 (MW = 49 kDa, red arrow). NaK-ATPase loading control (MW = 198 kDa, red arrow). (N = 1 donor, 2 replicates scr-gRNA control, 3 replicates KCNJ6 gRNAs, no virus control, and lentivirus KCNJ6 vector.) D, Representative images of isogenic control (iso-CTL) neurons, CRISPRa and Lenti expression ↑GIRK2 neurons stained for TUJ1 (magenta), DAPI (blue), and GIRK2 (green); 100× magnification; scale bar and insets = 10 μm. E, Normalization of GIRK2 puncta μm² to TUJ1 μm²; each point represents a single image; coral, CRISPRa and teal, Lenti [N images = CRISPRa: 9 iso-CTL (scr-gRNA control), 16 ↑GIRK2 (KCNJ6 gRNA); KCNJ6 lentivirus: 9 iso-CTL (no virus control), 10 ↑GIRK2]. All experiments were carried out in Day 42 NGN2 neurons.