Research Paper Volume 3, Issue 1 pp 33—43

Figure 2. HIF-1α regulates HIPK2 protein degradation. (A) HIF-1α-Flag (4, 6, and 8 μg) and HIPK2-GFP (4 μg) expression vectors were co-transfected in 293 cells where immunoblot analyses showed that increased amounts of HIF-1α induced HIPK2 abolishment. (B) Immunoblot analysis of C38 cells transfected with HIF-1α expression and treated with proteasome inhibitor MG132 (10 μM for 4 h) or DMSO vehicle. The endogenous HIPK2 levels downregulated by HIF-1α were rescued by MG132. (C) RT-PCR analysis of HIPK2 in RKO colon cancer cells co-transfected with HIF-1α and HIPK2 expression vector. (D) Immunoblot of endogenous HIPK2 protein in C38 and C27 prostate cancer cells, showing lower HIPK2 levels in “constitutive hypoxic” C27 cells. (E) The HIPK2 mRNA levels were comparable between C38 and C27 cells. (F) Immunoblot in C27 cells showed rescue of endogenous HIPK2 protein levels after HIF-1α knock-down by siRNA.