ZHX2 restricts hepatocellular carcinoma by suppressing stem cell-like traits through KDM2A-mediated H3K36 demethylation

Abstract

Background: Liver cancer stem cells (CSCs) are critical determinants of HCC relapse and therapeutic resistance, but the mechanisms underlying the maintenance of CSCs are poorly understood. We aimed to explore the role of tumor repressor Zinc-fingers and homeoboxes 2 (ZHX2) in liver CSCs. Methods: CD133 + or EPCAM + stem-like liver cancer cells were sorted from tumor tissues of HCC patients and HCC cell lines by flow cytometry. In addition, sorafenib-resistant cells, tumor-sphere forming cells and side population (SP) cells were respectively cultured and isolated as hepatic CSCs. The tumor-initiating and chemoresistance properties of ZHX2-overexpressing and ZHX2-knockdown cells were analyzed in vivo and in vitro. Microarray, luciferase reporter assay, chromatin immunoprecipitation (ChIP) and ChIP-on-chip analyses were performed to explore ZHX2 target genes. The expression of ZHX2 and its target gene were determined by quantitative RT-PCR, western blot, immunofluorescence and immunohistochemical staining in hepatoma cells and tumor and adjacent tissues from HCC patients. Results: ZHX2 expression was significantly reduced in liver CSCs from different origins. ZHX2 deficiency led to enhanced liver tumor progression and expansion of CSC populations in vitro and in vivo. Re-expression of ZHX2 restricted capabilities of hepatic CSCs in supporting tumor initiation, self-renewal and sorafenib-resistance. Mechanically, ZHX2 suppressed liver CSCs via inhibiting KDM2A-mediated demethylation of histone H3 lysine 36 (H3K36) at the promoter regions of stemness-associated transcription factors, such as NANOG, SOX4 and OCT4. Moreover, patients with lower expression of ZHX2 and higher expression of KDM2A in tumor tissues showed significantly poorer survival. Conclusion: ZHX2 counteracts stem cell traits through transcriptionally repressing KDM2A in HCC. Our data will aid in a better understanding of molecular mechanisms underlying HCC relapse and drug resistance.

Epigenetics in hepatocellular carcinoma development and therapy: The tip of the iceberg

“…For instance, increased KDM3A expression was associated with tumour recurrence after resection, 119 while the upregulation of LSD1/KDM1, KDM5B, KDM6B, KDM4B and KDM2A has been related to tumour aggressiveness and poor prognosis. [120] , [121] , [122] , [123] , [124] The transcriptional programmes influenced by the overexpression of KDMs are complex to elucidate. The activity of these enzymes may have differential effects on gene expression depending on the specific histone residue on which they act.…”

“…For example, LSD1/KDM1 and KDM4B, both upregulated in HCC, can remove repressive H3K9 methyl marks, while concomitantly, LSD1/KDM1 and KDM4B can eliminate H3K4 and H3K36 active methyl marks, respectively. 59 Nevertheless, several pro-tumorigenic mechanisms triggered by KDMs in HCC have been elucidated, including the promotion of stem cell-like traits by KDM6B and KDM2A, 122 , 124 and the maintenance of glycolytic metabolism, stemness and drug resistance by LSD1/KDM1. 125 , 126 …”

KDM2A Targets PFKFB3 for Ubiquitylation to Inhibit the Proliferation and Angiogenesis of Multiple Myeloma Cells

“…Studies have shown that KDM2A as a lysine demethylase was highly overexpressed in many cancers, such as breast cancer (8,29), gastric cancer (9), lung cancer (30) and hepatoma (31). In addition, it can be downregulated by tumor microenviroment (28).…”

ZHX2 inhibits SREBP1c‐mediated de novo lipogenesis in hepatocellular carcinoma via miR‐24‐3p

“…It is possible that ZHX2 might have other downstream targets than miR‐24‐3p. In fact, ZHX2 inhibits the development of HCC through many reported routes, such as inhibiting cell proliferation, the cell cycle, and cell migration [21,43]. Further efforts are needed to explore alternative mechanistic routes for ZHX2 inhibiting the development of HCC.…”