Figure 6: Hmga1 directly induces Sox9 expression.

(a) Relative Sox9 mRNA (mean±s.d.) by qPCR in GFP⁺ ISCs from WT and Hmga1 mice is shown from two experiments performed in triplicate. Gapdh was used to control for loading. **P<0.01; Student’s t-test. (b) Hmga1 binding to the Sox9 promoter by ChIP is shown from mouse crypt cells. IgG antibody and the Hprt promoter region were used as negative controls; histone H3 was a positive control. Bars show mean enrichment±s.d. from two experiments performed in triplicate; **P<0.01; two-tailed Student’s t-test. (c) Sox9 IHC staining (brown) is shown in intestinal cross-sections from WT and Hmga1 transgenic mice. Scale bars, 20 μm. (d) Western blots for Sox9, Hmga1, and Gapdh (loading control) were performed from freshly isolated crypt cells from WT and Hmga1 transgenic mice. Western blots were done three times; a representative blot is shown. Size markers (kDa) are indicated. (e) Densitometry analysis±s.d. was performed on repeat western blots. *P<0.05; two-tailed Student’s t-test. (f) Sox9 and Hmga1 IHC staining (brown) is shown in adjacent sections of small intestine from WT and Hmga1 mice. Arrows show representative staining for Hmga1 (H) or Sox9 (S); triangles show Hmga1 transgenic lymphocytes which have high levels of Hmga1 (but not Sox9). Scale bars, 50 μm. (g) Mean number of cells±s.d. staining positive for Hmga1, Sox9 or both were identified by IHC in adjacent cross-section on slides of small intestinal crypts from WT and Hmga1 mice (n≥21 crypts per group.) **P<0.01 by Mann–Whitney test.