Fig. 6

Med12 depletion reduces mNS-5 NSC proliferation likely through a G1/S phase cell cycle block. a-e mNS-5 NSCs were infected with lentiviruses expressing NS control, Med12-, Med1-, Cdk8-, and/or CycC (Ccnc)-specific shRNAs as indicated. a Five days post-infection, cells were harvested, fixed, and monitored for cell cycle distribution using a FACSCalibur flow cytometer and CellQuest Pro software prior to data analysis using FlowJo software. At least 10,000 single cell events were collected for the analysis. b Five days post-infection, cells were stained with annexin V conjugated to APC and propidium iodide. Stained cells were analyzed by FACSCalibur and data was analyzed using FlowJo software. c Lentivirus-infected cells were seeded at a concentration of 5 × 10⁴ cells/well in 12-well plates. Cell proliferation was monitored by counting viable cells using 0.4 % trypan blue at 24, 48, 72, and 96 h post-seeding. d, e mRNA expression levels for the indicated genes were determined by RT-qPCR analyses as described in Fig. 2. mRNA levels for each gene were normalized to Gapdh mRNA and expressed relative to their corresponding mRNA levels in control NS shRNA-expressing cells. All data represent the mean +/− SEM of at least three independent experiments performed in triplicate. p values were calculated by Student’s t test. Asterisks denote statistically significant differences relative to NS control shRNA (*p < 0.1, **p < 0.05, ***p < 0.01)