Figure 2

CHD7 expression in freshly dissected tumor tissue and in human-glioblastoma derived cell lines. (A) Relative CHD7 mRNA levels of freshly dissociated CD133^pos and CD133^neg tumor cells were assessed by qRT-PCR. Cell fractions represent matched sub-populations from the same patient. Results are expressed as average ± SEM for technical replicates. ^***p < 0.001; 2-way ANOVA followed by Bonferroni posttest. (B) Relative CHD7 mRNA levels in different LTCs and GICs, determined by qRT-PCR. The results represent average ± SEM from two independent experiments. ^##p < 0.01, ^***p < 0.001; one-way ANOVA followed by Bonferroni correction for multiple tests compared with LN-229 and all the other cell lines, respectively. (C) CHD7 immunoblotting of fractionated nuclear extracts (NE) and cytoplasmic extracts (CE) of LN-229 and LN-319 cell lines. PARP1 and HSP90 were used as nuclear and cytoplasmic markers, respectively, examined consecutively in the same blot as CHD7, after membrane stripping. (D) CHD7 immunoblotting of nuclear extracts of human glioblastoma-derived cell lines. Left panel shows the result from LTCs. In a separate gel, right panel, is the blot from the GICs. Actin was used as loading control for each of the gels. Due to the great difference in the protein sizes, the loading control was examined in a separate gel, loaded under the same conditions. Total protein extract of 293 T cells transfected with the empty vector and CHD7 overexpression plasmids were used as negative and positive controls, respectively.