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. 2019 Apr 12;20(6):e47227. doi: 10.15252/embr.201847227

Figure 5. cTAZ expression was induced by JAK‐STAT signaling.

Figure 5

  1. gVSV induced expression of cTAZ. RKO cells were infected with gVSV (MOI = 0, 0.001, 0.01, 0.01, 1) for 8 h. Protein expression was determined by IB (Left) and quantified (Right). Error bars indicate SD, n = 3. ***P < 0.001; two‐way ANOVA test was used for statistical analysis.
  2. IFN‐α induced expression of cTAZ. RKO cells were treated with different doses of IFN‐α for 8 h. Protein expression was determined as in (A). Error bars indicate SD, n = 3. ***P < 0.001; two‐way ANOVA test was used for statistical analysis.
  3. IFN‐α induced mRNA level of cTAZ. RKO cells were treated with different doses of IFN‐α for 8 h. RNA levels were measured by qPCR. Error bars indicate SD, n = 3. ***P < 0.001; two‐way ANOVA test was used for statistical analysis.
  4. STAT1/2 synergistically activated a cTAZ promoter. HEK293A cells were transfected with cTAZ reporter with or without STAT proteins. Promoter activity was determined using luciferase assay, and protein expression was determined by IB. Error bars indicate SD, n = 3. ***P < 0.001; Student's t‐test.
  5. STAT1/2 binding sites on cTAZ promoter. HEK293A cells were transfected with cTAZ promoters (wild type or mutants). Luciferase activity and responses to STAT1/2 expression were measured. Error bars indicate SD, n = 3. **P < 0.01; Student's t‐test.
  6. STAT1 occupied cTAZ promoter in an IFN‐α‐sensitive manner. RKO cells were treated with or without IFN‐α (50 ng/ml) for 1 h and subjected to ChIP assays. cTAZ PCR primers were designed to target STAT1/2 binding site. ISG15 promoter was included as a positive control.
  7. A negative feedback mechanism regulating JAK‐STAT signaling involving cTAZ. Upon IFN‐α stimulation, STAT1/2 are phosphorylated and undergo dimerization. STAT1/2 dimer interacts with importins (such as Im‐α5), enters nucleus, and activates transcription of target genes including ISGs. The expression of cTAZ is induced by STAT1/2. Elevated cTAZ blocks the dimerization and nuclear translocation of STAT1/2, which in turn leads to repression of the JAK‐STAT signaling. cTAZ represents a critical node of a negative feedback mechanism regulating the JAK‐STAT signaling.

Source data are available online for this figure.