Fig. 7: Reduction of 5hmC causes a phenotype switch of BSMCs.
From: Spurious transcription causing innate immune responses is prevented by 5-hydroxymethylcytosine
a, EM images of control and Tet3smKO bronchial SMCs (n = 4). Arrows point to dense bodies (control) and rough endoplasmic reticulum (Tet3smKO). Scale bar, 1,000 nm. b, Western blot analysis of sorted lung SMCs. Pan-actin was used as loading control. c, Western blot analysis of mESC-SMCs after transduction with scramble, Tet3KD, Tet3KD + TET3WT and Tet3KD + TET3CD lentiviruses. Pan-actin was used as loading control. Quantification of protein levels is shown on the right (n = 3 independent experiments; one-way ANOVA with Tukeyʼs post hoc test: *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001). d, RT-qPCR analysis of indicated genes in mESC-SMCs after transduction with scramble, Tet3KD, Tet3KD + TET3WT, Tet3KD + TET3CD lentiviruses (n = 4 independent experiments; one-way ANOVA with Tukey’s post hoc test: *P < 0.05; **P < 0.01; ***P < 0.001). e, ChIP–qPCR to monitor Pol II pSer5 enrichment within gene bodies of indicated genes in mESC-SMCs after transduction with scramble, Tet3KD, Tet3KD + TET3WT, Tet3KD + TET3CD lentiviruses with DRB treatment (n = 4 independent experiments; one-way ANOVA with Tukeyʼs post hoc test: *P < 0.05; **P < 0.01; ***P < 0.001). f, Semiquantitative RT-PCR analysis of Acta2, Cnn1, Dbn1, Vim and Tpm4 full-length mRNA in mESC-SMCs after transduction with scramble, Tet3KD, Tet3KD + TET3WT and Tet3KD + TET3CD lentiviruses. β-actin was used for normalization. Quantification was performed by Image J and is shown on the right (n = 3 independent experiments; one-way ANOVA with Tukeyʼs post hoc test: *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001). Data in c–f are presented as mean values ± s.e.m.
