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Fig. 4 | Journal of Experimental & Clinical Cancer Research

Fig. 4

From: The JAK2/STAT3/CCND2 Axis promotes colorectal Cancer stem cell persistence and radioresistance

Fig. 4

CCND2 is a downstream target of JAK2/STAT3 signaling. a Potential upstream regulators were analyzed in residual CRC tissues after RT (GSE 15781) using IPA software. The left Y-scale indicates enrichment z-score, the size of the round symbol indicates the ratio of contributing components, and the color indicates the p-value. The right panel shows the JAK2 networks interacting with other upstream regulators. b The potential downstream target genes of JAK2 in residual CRC tissues after RT (GSE 15781) were validated in vitro using shCTRL- or shJAK2-transfected HCT116 cells cultured in monolayer or stem cell-enriched sphere-forming conditions. c-e The expression of JAK2 and CCND2 was measured in CRC patient-derived tumor tissues and adjacent normal tissues at (c) mRNA and (d) protein levels. e Based on the mRNA expression level, a significant positive correlation between JAK2 and CCND2 was observed by Pearson correlation analysis. f In CRC patient tumors obtained from R2 database (https://hgserver1.amc.nl/cgi-bin/r2/main.cgi, GSE41258 and GSE37892), a significant positive correlation between JAK2 and CCND2 was identified by the same method above. g and h The radiation-induced expression of CCND2 was assessed by Western blot at 48 h after radiation under JAK2 knockdown or Stattic treatment in (g) HCT116 or (h) LoVo cells. i Transcriptional activity of STAT3 on the CCND2 promoter was confirmed by the luciferase assay with or without Stattic treatment. j Predicted binding sites of STAT3 on the CCND2 promoter region by the Champion ChIP Transcription Factor Search Portal (Qiagen); accordingly, primers for the ChIP assay were designed. k The ChIP assay was conducted in HCT116 cells with or without Stattic treatment. Immunoprecipitation was conducted using STAT3- and STAT3-bound DNA fragments amplified by PCR. PCR products were confirmed by gel electrophoresis. The bar graph represents the mean ± SD (n = 3). Statistical analyses were implemented using one-way ANOVA followed by Dunnett’s multiple comparison test against the control group or by Student’s t-test between two groups; *, **, and *** indicate p < 0.05, p < 0.01, and p < 0.001, respectively

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