RYBP Expression Is Regulated by KLF4 and Sp1 and Is Related to Hepatocellular Carcinoma Prognosis

Qiaojiajie Zhao,Weihua Cai,Xuan Zhang,Shuo Tian,Junwen Zhang,Haibo Li,Congcong Hou,Xiaoli Ma,Hong Chen,Bingren Huang,Deng Chen

doi:10.1074/jbc.m116.770727

Abstract

The expression of Ring1- and YY1-binding protein (RYBP) is reduced in several human cancers, but the molecular mechanism(s) have remained elusive. In this study, we used human hepatocellular carcinoma (HCC) cell lines and tissue specimens to study the mechanism and herein report several new findings. First, we cloned and characterized the basal promoter region of the human RYBP gene. We found that the decreased RYBP expression in HCC tissues was not due to promoter sequence variation/polymorphisms or CpG dinucleotide methylation. We identified two transcription factors, KLF4 and Sp1, which directly bind the promoter region of RYBP to induce and suppress RYBP transcription, respectively. We mapped the binding sites of KLF4 and Sp1 on the RYBP promoter. Studies in vitro showed that KLF4 suppresses whereas Sp1 promotes HCC cell growth through modulating RYBP expression. Deregulated KLF4 and Sp1 contributed to decreased expression of RYBP in HCC tumor tissues. Our studies of human HCC tissues indicated that a diminished RYBP level in the tumor (in association with altered KLF4 and Sp1 expression) was statistically associated with a larger tumor size, poorer differentiation, and an increased susceptibility to distant metastasis. These findings help to clarify why RYBP is decreased in HCC and indicate that deregulated KLF4, Sp1, and RYBP may lead to a poorer prognosis. Our findings support the idea that RYBP may represent a target for cancer therapy and suggest that it may be useful as a prognostic biomarker for HCC, either alone or in combination with KLF4 and Sp1.

Highlights

The expression of Ring1- and YY1-binding protein (RYBP) is reduced in several human cancers, but the molecular mechanism(s) have remained elusive
The RYBP Expression in hepatocellular carcinoma (HCC) Cell Lines, the Cloning and Characterization of the Full Promoter Region, and the Lack of Importance of CpG Methylation and Promoter Variations/Polymorphisms on the RYBP Expression—Previous studies have shown that the expression of RYBP mRNA and protein is
To elucidate the molecular mechanism underlying the reduced expression, we first examined the mRNA levels of RYBP in immortalized human hepatocyte (IHH) and five HCC cell lines (Huh7, HepG2, Hep3B, PLC/PRF/5, and SK-Hep-1), which we intended to use as cellular models

Summary

Introduction

The expression of Ring1- and YY1-binding protein (RYBP) is reduced in several human cancers, but the molecular mechanism(s) have remained elusive. Studies in vitro showed that KLF4 suppresses whereas Sp1 promotes HCC cell growth through modulating RYBP expression. Our studies of human HCC tissues indicated that a diminished RYBP level in the tumor (in association with altered KLF4 and Sp1 expression) was statistically associated with a larger tumor size, poorer differentiation, and an increased susceptibility to distant metastasis. In agreement with its apoptosis-inducing capacity, the expression of RYBP has been reported to be reduced in a variety of human cancers, including lung, cervical, prostate, and liver cancers, and was recently shown to inhibit cancer growth, metastasis, and chemoresistance in vivo and in vitro [14,15,16,17], indicating that it is a potential candidate drug target for use against these tumors. We demonstrate that the deregulation of KLF4, Sp1, and RYBP is related to a more malignant phenotype of HCC

Methods

Results

Conclusion