Fig. 7

MYH1G-AS reduces interaction of FGF18 to SMARCA5, thereby promoting the transcription and expression of SMAD4. A The binding of FGF18 to the SMARCA5 protein was determined by Co-IP assay. B The binding of SMARCA5 to the FGF18 protein was determined by Co-IP assay. C The interaction between FGF18 and the SMARCA5 protein was determined by the yeast two-hybrid system. D IF staining of SMARCA5 and FGF18 in CPM. E The interaction between FGF18 and the SMARCA5 protein was determined by Co-IP assay. F Co-IP assay after cotransfection with pcDNA3.1-FGF18-FLAG and the listed nucleic acids. (G) Relative SMAD4 expression after MYH1G-AS interference detected by RNA-seq. H−K Relative mRNA (H and J) and protein (I and K) expression levels of SMAD4 with MYH1G-AS overexpression or knockdown in vitro (H, I) and in vivo (J−K). L Left: Schematic of four truncated SMAD4 promoter constructs used for luciferase assays. Right: Dual-luciferase reporter assays of four reporter constructs. M The transcriptional activity of SMAD4 core promoter region. N ChIP analysis of the binding capacity of POU2F1 to the SMAD4 promoter. O, P Relative POU2F1 (O) and SMAD4 (P) expression after POU2F1 overexpression or knockdown. Q The transcriptional activity of the SMAD4 core promoter region after MYH1G-AS overexpression or knockdown. R ChIP analysis of the binding capacity of POU2F1 to the SMAD4 promoter with MYH1G-AS overexpression or knockdown. In panels F, I, and K, the numbers shown below the bands were folds of band intensities relative to control. Band intensities were quantified by ImageJ and normalized to β-tubulin. Results are shown as mean ± SEM. In panels G, H, J, and L− R, the statistical significance of differences between means was assessed using an independent sample t-test. (**P < 0.01)