Figure 7. ZNF277 deficiency augments p21^WAF1 expression.

(A) siRNA and CRISPR knockdown of ZNF277 expression augments p21^WAF1 levels in HT29, H508, and HEK293 cells. β-Actin was used as a loading control. (B) Levels of murine p21^WAF1 expression are augmented in colon tissue extracts from Apc^Min/+ and Zfp277^–/– mice. Experiments were performed using tissues from 2 separate 8-week-old mice of each genotype. (C) Upregulated p21^WAF1 expression after p53 and ZNF277 knockdown in HT29 cells. All siRNAs were 25 nM, except lane 4 (50 nM). (D) ZNF277 knockdown augments p21^WAF1 mRNA levels in HT29 cells. Data are shown as mean ± SEM from 3 separate experiments. *P < 0.01 (2-tailed Student’s t test). (E) Zfp277 deficiency stimulates cellular senescence. β-Galactosidase staining in control HT29 cells (A) and HT29 cells with CRISPR knockdown of ZNF277 (B). Scale bar: 50 μM. (F) Zfp277 deficiency increases p21^WAF1 expression. IHC of p21^WAF1 in the normal small intestine of WT (A) and Zfp277^–/– (B) mice, as well as in small intestine adenomas from Zfp277^+/+Apc^Min/+ (C) and Zfp277^–/–Apc^Min/+ (D) mice. Arrows indicate adenomas. Scale bar: 100 μM.