UHRF1 Regulates the Protein Level of HIF-1α Dependents on Its Own Methylation Modification
WANG Di, GAO Chao, WANG Mengdong, XIAO Chenglu*, TANG Jun*
HIF-1 (hypoxia-inducible factor-1) plays a key role in hypoxia response system. HIF-1 is a heterodimer transcriptional factor formed by HIF-1α and HIF-1β. This study aimed to explore the effect of UHRF1 (ubiquitin-like with PHD and RING finger domains 1) on HIF-1α protein under hypoxic conditions. UHRF1- targeted small interfering RNA was used to inhibit the expression of UHRF1. Western blot and qRT-PCR were used to detect the expression levels of HIF-1α protein and mRNA respectively. UHRF1 was overexpressed in HeLa and HepG2 cells, and the protein expression level of HIF-1α was detected by Western blot. SET7/9 (SET domain containing histone lysine methyltransferase 7/9) and UHRF1 were overexpressed or knocked down synchronously, and the expression level of HIF-1α protein was detected by Western blot. The interaction between full length UHRF1 or different truncated UHRF1 proteins and HIF-1α protein was detected by co-immunoprecipitation. The 385th lysine of UHRF1 was mutated to arginine, and the interaction between the mutant protein and HIF-1α protein was detected by co-immunoprecipitation. The results showed that inhibition of UHRF1 expression could upregulate the protein level of HIF-1α under hypoxic conditions, but had no effect on the mRNA level of HIF-1α. UHRF1 overexpression didn’t affect HIF-1α protein level. However, when UHRF1 and SET7/9 were overexpressed synchronously, the protein level of HIF-1α decreased. In addition, simultaneous inhibition of UHRF1 and SET7/9 expression could rescue the increased HIF-1α protein level when UHRF1 expression was inhibited. It was further found that UHRF1 and HIF-1α could interact with each other, and this interaction depended on the 385th lysine methyl modification of UHRF1, and the methylation modification was regulated by SET7/9. SET7/9 mediated methyl modification of UHRF1 is necessary for its interaction with HIF-1α and its effect on the protein level of HIF-1α.
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