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. 2016 Jun 16;7:11798. doi: 10.1038/ncomms11798

Figure 6. PP2A regulates STAT3 activation at S727 and mediates suspension survival in TICs.

Figure 6

(a) Bulk cancer cells (Bulk) and spheroid culture-enriched TICs (SPH) were cultured under spheroid condition for 10 h, followed by western blot assay. (b) Spheroid culture of PKH-labelled HT29 cells was assayed for immunostaining, and fluorescence intensity was quantified with ZEISS microscope software ZAN. (c) Western blot analysis for CCS bulk cancer cells treated with 0, 0.1, 0.5 and 1 nM OA for 1 h or 0, 5, 10 and 15 nM CA for 30 min. (d) Western blot analysis for CCS spheroid culture-enriched TICs treated with 50 nM ceramide C6 for 24 h. (e) Western blot analysis for immunoprecipitated PP2A or STAT3 complex of CCS bulk cancer cells. (f) Spheroid culture-enriched TICs expressing control plasmid (CTR) or WT PP2A were subjected to spheroid condition for 24 h. Top, western blot analysis. Bottom, quantification of TUNEL-positive cells. (g) Bulk cancer cells expressing control plasmid (CTR) or DN PP2A were subjected to spheroid condition for 24 h. Top, western blot analysis. Bottom, quantification of TUNEL-positive cells. (h) Cells in f were subjected to in vivo suspension condition for 24 h, followed by TUNEL assay. (i) Cells in g were subjected to in vivo suspension condition for 24 h, followed by TUNEL assay. The results are expressed as mean±s.d. of three independent experiments. Asterisks indicate significant differences (**P<0.01) determined by Student's t-test. Scale bar, 20 μm.