A positive-feedback loop between HBx and ALKBH5 promotes hepatocellular carcinogenesis

Abstract

Background Hepatitis B Virus (HBV) contributes to liver carcinogenesis via various epigenetic mechanisms. The newly defined epigenetics, epitranscriptomics regulation, has been reported to involve in multiple cancers including Hepatocellular Carcinoma (HCC). Our previous study found that HBx, HBV encodes X protein, mediated H3K4me3 modification in WDR5-dependent manner to involve in HBV infection and contribute to oncogene expression. AlkB Homolog 5 (ALKBH5), one of epitranscriptomics enzymes, has been identified to be associated with various cancers. However, whether and how ALKBH5 is dysregulated in HBV-related HCC remains unclear yet. This study aims to investigate ALKBH5 function, clinical significance and mechanism in HBV related HCC (HBV-HCC) patients derived from Chinese people. Methods The expression pattern of ALKBH5 was evaluated by RT-qPCR, Western blot, data mining and immunohistochemistry in total of 373 HBV-HCC tissues and four HCC cell lines. Cell Counting Kit 8 (CCK8) assay, Transwell and nude mouse model were performed to assess ALKBH5 function by both small interference RNAs and lentiviral particles. The regulation mechanism of ALKBH5 was determined in HBx and WDR5 knockdown cells by CHIP-qPCR. The role of ALKBH5 in HBx mRNA N6-methyladenosine (m6A) modification was further evaluated by MeRIP-qPCR and Actinomycin D inhibitor experiment in HBV-driven cells and HBx overexpression cells. Result ALKBH5 increased in tumor tissues and predicts a poor prognosis of HBV-HCC. Mechanically, the highly expressed ALKBH5 is induced by HBx-mediated H3K4me3 modification of ALKBH5 gene promoter in a WDR5-dependent manner after HBV infection. The increased ALKBH5 protein catalyzes the m6A demethylation of HBx mRNA, thus stabilizing and favoring a higher HBx expression level. Furthermore, there are positive correlations between HBx and ALKBH5 in HBV-HCC tissues, and depletion of ALKBH5 significantly inhibits HBV-driven tumor cells’ growth and migration in vitro and in vivo. Conclusions HBx-ALKBH5 may form a positive-feedback loop to involve in the HBV-induced liver carcinogenesis, and targeting the loop at ALKBH5 may provide a potential way for HBV-HCC treatment.

ALKBH5/MAP3K8 axis regulates PD-L1+ macrophage infiltration and promotes hepatocellular carcinoma progression

“…Recently, emerging evidence has suggested that abnormal signal pathway activation, tumor microenvironment formation, and immunosuppressive states are involved in the occurrence and development of HCC. Although a few studies have reported ALKBH5 involvement in the pathogenesis of HCC, their results have been contradictory 10 , 11 , 49 . This may be attributed to the selection of tumor cell lines, the efficiency of virus transfection and experimental conditions.…”

“…Besides, Depmap data based on CRISPR data also showed that silencing ALKBH5 impaired the proliferation of hepatoma cells including Huh-7. In two studies supporting ALKBH5 to promote liver cancer, only 2 liver cancer cell lines (HepG2 and MHCC-97H) were used, and only knockdown of ALKBH5 was done 10 , 49 . We used several authoritative liver cancer data sets and collected clinical samples to verify the high expression of ALKBH5 in liver cancer and its correlation with poor prognosis.…”

The bioinformatics and experimental analysis of AlkB family for prognosis and immune cell infiltration in hepatocellular carcinoma

“…There are many findings indicated that m6A modification mediated by ALKBH5 and FTO participates in the development of HCC and plays a crucial role in suppressing proliferative and invasive abilities of HCC cells ( Bian et al, 2021 ; Chen et al, 2020 ). The expression of ALKBH5 and FTO has been proved to be up-regulated in tumor tissues, which implies a poor prognosis of HCC patients ( Li et al, 2019 ; Qu et al, 2021 ). Apart from that, m6A is evidently connected with HCC immune microenvironment.…”

Role of m6A writers, erasers and readers in cancer

“…ALKBH5 plays a dual role as a carcinogen and tumor suppressor in different cancers and, in certain cases, in the same type of cancer. ALKBH5 promotes cancer progression by regulating TIMP3 [ 105 ], FOXM1 [ 106 ], CDKN1A [ 107 ], JAK2 [ 108 ], FOXM1 [ 101 , 109 , 110 ], AURKB [ 111 ], G6PD [ 112 ], HBx [ 113 ], USP1 [ 114 ], NANOG [ 115 ], PVT1 [ 116 ], IGF1R [ 117 ] lncRNA NEAT1 [ 118 , 119 ], and lncRNA RMRP [ 120 ] expression. In addition, ALKBH5 inhibited cancer progression by regulating PD-L1 [ 103 ], CK2α [ 121 ], LYPD1 [ 102 ], PER1 [ 122 ], WIF-1 [ 99 ], and lncRNA KCNK15-AS1 [ 123 ] expression.…”