Fig. 4.

NIPBL loss inhibits TYMS transcription through downregulating MYC bioactivity. (a) Mutation status of cohesin complex and -associated members in TCGA provisional stomach database. 395 tumour samples are available for analysing the mutation status of cohesin complex members. (b) The protein expression of NIPBL, MYC and thymidylate synthase were detected in multiple gastrointestinal cancer cell lines. (c) Gene Set Enrichment Analysis of NIPBL knockdown versus scramble in NUGC3 cells. (d) Indicated proteins were detected within 7 days using immunoblotting after inducible knockdown of NIPBL. (e) Indicated proteins were detected by immunoblotting after NIPBL was knocked down in another three gastrointestinal cancer cell lines. (f) The MYC and TYMS mRNA expression levels were detected by real-time PCR in NUGC3 cells after NIPBL knockdown, normalized with GAPDH mRNA expression levels. (g) The luciferase activity of TYMS promoter was detected in NUGC3 cells after NIPBL knockdown. (h) The luciferase activity of MYC promoter (−3,255 to −1,719 from ATG site) was detected in HCT116 cells after NIPBL knockout. (i) The luciferase activity of TYMS promoter was detected after MYC was overexpressed in NIPBL knockout HCT116 cells. Data are shown as mean ± SD. *p < 0.05, **p < 0.01, ***p < 0.001, p value was calculated with by two-tailed Student's t-test.