Loss of GATA 5 expression due to gene promoter methylation induces growth and colony formation of hepatocellular carcinoma cells

GATA5 is a transcription factor that is capable of suppressing the development of various types of human cancer. The present study investigated the expression of GATA5 and GATA4, and examined their roles in the proliferation and colony formation ability of hepatocellular carcinoma (HCC) tissues and cells. The GATA4 and GATA5 expression levels and gene promoter methylation of HCC tissue samples from 38 patients and HCC cell lines were analyzed using reverse transcription-polymerase chain reaction (RT-PCR) and methylation‐specific PCR (MSP), respectively. The effects of GATA4 and GATA5 overexpression on the proliferation and colony forming ability of HCC cells were also assessed using cell viability and colony formation assays. A luciferase reporter assay was utilized to investigate the transcriptional interaction of GATA4 and GATA5 with canonical Wnt signaling. The results indicated that the expression levels of GATA4 and GATA5 were lost or reduced following methylation of gene promoters in HCC tissues and cell lines. Treatment with a demethylating agent, 5-aza-2'-deoxycytidine (5-AZA), restored GATA4 and GATA5 expression in HCC cell lines. Furthermore, methylation of the GATA5 promoter was observed to be associated with the age of patients exhibiting HCC. Restoration of GATA4 and GATA5 expression inhibited colony formation and induced apoptosis of HCC cells in vitro. The present study concluded that the expression levels of GATA4 and GATA5 were reduced in HCC tissues and cell lines. Treatment with 5-AZA restored GATA4 and GATA5 expression in HCC cell lines, suppressing tumor cell growth and colony formation, as well as inducing apoptosis. Introduction Primary liver cancer is the fifth most common cause of cancer-associated mortality worldwide (1), and up to 90% of primary liver cancers are hepatocellular carcinomas (HCC) (2). Risk factors for hepatocarcinogenesis include chronic hepatitis B or C infection, alcohol consumption and aflatoxin B1 contamination (2). HCC development, similarly to that of other types of cancer, is a multistep process, involving numerous genetic and epigenetic alterations (3). It has been reported that promoter methylation of certain tumor suppressor genes occurs frequently during HCC development and progression (4,5). The GATA gene family comprises a group of zinc finger transcription factors that are able to bind to the GATA motif (WGATAR), which is present in the promoters of certain genes (6). GATA1, GATA2 and GATA3 are functionally involved in cellular lineage determination (7), while GATA4, GATA5 and GATA6 regulate the development of endoderm-derived organs, including the heart and gut (8). During development, GATA4 and GATA5 induce differentiation of embryonic stem cells into specific, mature gut cells (9). In adults, GATA4 and GATA5 regulate epithelial cell differentiation (10,11). However, altered expression of GATA4 and GATA5 proteins is associated with tumorigenesis in certain organs and tissues, including gastric and colon cancer; thus, GATA4 and GATA5 may act as putative tumor suppressor genes (11,12). A previous study revealed that GATA4 and GATA5 expression was lost in ovarian and gastric cancer, and demonstrated that chromosomal regions of GATA4 (8p23.1-p22) and GATA5 (20q13.2-q13.3) loci were frequently deleted in various types of human cancer (13). Additional studies identified that loss of GATA4 and GATA5 expression during human neoplastic progression, including that of pancreatic, non-small cell lung, esophageal and renal cancer, was due to promoter methylation (14-16). Loss of GATA4 and GATA5 expression may also alter the typical expression patterns of numerous downstream gene networks with antineoplastic properties (11). For example, GATA transcription factors are able to integrate Wnt signaling during heart muscle formation (17,18) and alteration of the Wnt/β-catenin pathway has a critical role in the development of various types of human cancer, including HCC (19). Loss of GATA5 expression due to gene promoter methylation induces growth and colony formation of hepatocellular carcinoma cells LEI XIA1*, YAN GONG2*, AIQUN ZHANG3, SHOUWANG CAI3 and QIANG ZENG2 1Department of Medical Security; 2Health Management Institute; 3Department of Hepatobiliary Surgery, Chinese People's Liberation Army General Hospital, Beijing 100853, P.R. China Received October 24, 2014; Accepted August 18, 2015 DOI: 10.3892/ol.2015. 3974 Correspondence to: Professor Qiang Zeng, Health Management Institute, Chinese People's Liberation Army General Hospital, 28 Fuxing Road, Beijing 100853, P.R. China E-mail: zq301@hotmail.com *Contributed equally