Figure 3

sh Fmr1 virus silenced FMRP expression and reduced cellular viability in ETO treated HeLa cells. To understand the role of FMRP on cellular viability, FMRP expression was inhibited by small hairpin RNA virus (shFmr1 lentivirus) as described in methods. RT-PCR band (A: top) and Real time PCR quantification (A: bottom) showed decreased level of Fmr1 mRNA. GAPDH was used as control. (B) Western blot assays was performed for FMRP and β-actin. (C) Apoptotic cell death after FMRP knock down. After shFmr1 virus-induced inhibition of FMRP, HeLa cells were treated with ETO for 3 hr and then propodium iodide (PI) staining was performed. Red fluorescence represents PI positive cells. (D) Bar graph represents the quantification of the PI-positive apoptotic cells. (E) After an appropriate treatment as above, cells were stained with TUNEL as described. (F) Graph represents the quantification of the apoptotic cell number. (G) Histograms show the results of FACS analysis. (H) The graph represents the quantification of the apoptotic cells in FACS analysis. Data represent mean ± S.E.M. ^# Significantly different as compared with control (p < 0.01, n = 4), ^# significantly different as compared with control virus (CV) group (p < 0.01, n = 4). Con: control, CV: sh non targeted control virus and FV: shFmr1 virus.