Journal: Proceedings of the National Academy of Sciences of the United States of America
Article Title: T-cell factor 3 (Tcf3) deletion increases somatic cell reprogramming by inducing epigenome modifications
doi: 10.1073/pnas.1017402108
Figure Lengend Snippet: NPC epigenome modifications induced by shTcf3. (A) Experimental scheme: NPCs-Oct4-puro were infected with OKshTcf3 or OKshCtrl. After 3, 5, and 7 d in NPC medium, the cells were trypsinized and replated in ES cell medium for an additional 24 h. AcH3 and H3K9me3 staining was then performed. (B) Representative images and quantification (Lower) of total H3 acetylation levels in NPCs infected with OKshCtrl or OKshTcf3. Intensity of AcH3 was evaluated using ImageJ software (mean ± SEM, n = 3; 100 cells in total were analyzed for each sample). (C) Representative images and quantification (Lower) of H3K9me3 heterochromatin foci. H3K9me3 staining was analyzed by immunofluorescence. The number of immunopositive H3K9me3 foci in the nuclei was counted (total of 70 nuclei were analyzed for each sample). Smoothing splines were fitted to the experimental data using the curve-fitting toolbox implemented in MATLAB R2009a (Mathworks). (D) Representative Western blot analysis of histone modifications of protein extracts from NPCs-Oct4-puro infected with OKshTcf3 or OKshCtrl. (E) Nucleus areas of OKshCtrl- and of OKshTcf3-infected NPCs. The cells were infected with OKshCtrl or OKshTcf3 and maintained in NPC medium for 7 d. Nucleus area was analyzed 24, 48, and 72 h after the shift to ES cell medium (mean ± SEM, n = 3; 100 cells were analyzed for each treatment time point and experiment; two-sample Wilcoxon test was used to calculate the P value).
Article Snippet: Smoothing splines were fitted to the experimental data using the curve-fitting toolbox implemented in MATLAB R2009a (Mathworks). ( D ) Representative Western blot analysis of histone modifications of protein extracts from NPCs-Oct4-puro infected with OKshTcf3 or OKshCtrl. ( E ) Nucleus areas of OKshCtrl- and of OKshTcf3-infected NPCs.
Techniques: Infection, Staining, Software, Immunofluorescence, Western Blot
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