Abstract 5328: RNF12 inhibits the proliferation of hepatocellular carcinoma through c-Myc and p21

Abstract

Abstract Purpose: RNF12/RLIM, a RING E3 ubiquitin ligase, plays an important role in the epigenetic regulation of sex-specific nurturing tissues in female mice. Recently, a few studies demonstrated a correlation between RNF12 and the proliferation of multiple cancers. The role of RNF12 in hepatocellular carcinoma is incompletely understood. The purpose of this study is to investigate the effect of RNF12 on the proliferation of human hepatocellular carcinoma cells. RNF12 is also known to interact with a number of nuclear proteins, including CLIM, HDAC2, and Smad4. We also explored whether c-Myc and p21, an oncogene and tumor suppressor essential to liver tumorigenesis, were affected by RNF12. Methods: Immunohistochemistry of RNF12 and c-Myc was performed on surgically dissected human hepatocellular carcinoma tissues. The soft-agar cloning, flow cytometry and Transwell invasion assays were carried out to investigate the role of RNF12 in SK-Hep1 cell proliferation, cell cycle and invasiveness. Real-time quantitative PCR and Western blotting were used to examine the mRNA and protein levels of RNF12 and related proteins in SK-Hep1 cells. Results: We found that the level of RNF12 was lower in hepatocellular carcinoma, compared with the surrounding non-cancerous tissues, while the c-Myc expression was higher. Using a SK-Hep1 cell line stably expressing RNF12, we showed that RNF12 over-expression inhibited cell proliferation and induced cell cycle arrest at G1 phase. Furthermore, we showed that RNF12 over-expression decreased the level of oncoprotein c-Myc, while not much affecting its mRNA level. Co-immunoprecipitation demonstrated that RNF12 and c-Myc associated with each other, suggesting a direct protein-protein interaction. RNF12 over-expression also markedly increased the mRNA and protein levels of p21, a tumor suppressor and the downstream target of p53, c-Myc and many other cancer-related regulators. Consistently, silencing of RNF12 reduced p21 mRNA and protein levels and increased c-Myc protein. Conclusion: Our study demonstrated reduced expression of RNF12 in hepatocellular carcinoma, and RNF12 over-expression inhibited the proliferation and cell cycle progression of SK-Hep1 cells. Mechanistically, RNF12 may directly regulate the level of c-Myc, similarly as other transcription factors such as CLIM and Smad4. The mechanisms underlying the increased expression of p21 is being investigated. Taken together, RNF12 may serve as a tumor suppressor in hepatocellular carcinoma, partly through oncoprotein c-Myc and tumor suppressor p21. Note: This abstract was not presented at the meeting. Citation Format: Meng Nie, Yongsheng Huang, Lin Wang. RNF12 inhibits the proliferation of hepatocellular carcinoma through c-Myc and p21 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 5328. doi:10.1158/1538-7445.AM2017-5328