Hepatitis B Virus–Telomerase Reverse Transcriptase Promoter Integration Harnesses Host ELF4, Resulting in Telomerase Reverse Transcriptase Gene Transcription in Hepatocellular Carcinoma

Abstract

Background and Aims Hepatitis B virus (HBV) integrations are common in hepatocellular carcinoma (HCC). In particular, alterations of the telomerase reverse transcriptase (TERT) gene by HBV integrations are frequent; however, the molecular mechanism and functional consequence underlying TERT HBV integration are unclear. Approach and Results We adopted a targeted sequencing strategy to survey HBV integrations in human HBV‐associated HCCs (n = 95). HBV integration at the TERT promoter was frequent (35.8%, n = 34/95) in HCC tumors and was associated with increased TERT mRNA expression and more aggressive tumor behavior. To investigate the functional importance of various integrated HBV components, we employed different luciferase reporter constructs and found that HBV enhancer I (EnhI) was the key viral component leading to TERT activation on integration at the TERT promoter. In addition, the orientation of the HBV integration at the TERT promoter further modulated the degree of TERT transcription activation in HCC cell lines and patients’ HCCs. Furthermore, we performed array‐based small interfering RNA library functional screening to interrogate the potential major transcription factors that physically interacted with HBV and investigated the cis‐activation of host TERT gene transcription on viral integration. We identified a molecular mechanism of TERT activation through the E74 like ETS transcription factor 4 (ELF4), which normally could drive HBV gene transcription. ELF4 bound to the chimeric HBV EnhI at the TERT promoter, resulting in telomerase activation. Stable knockdown of ELF4 significantly reduced the TERT expression and sphere‐forming ability in HCC cells. Conclusions Our results reveal a cis‐activating mechanism harnessing host ELF4 and HBV integrated at the TERT promoter and uncover how TERT HBV‐integrated HCCs may achieve TERT activation in hepatocarcinogenesis.

The Mechanisms of HBV-Induced Hepatocellular Carcinoma

“…36 Recent studies have shown that the binding of E74-like ETS transcription factor 4 (ELF4) to chimeric HBV EnhI in the TERT promoter region is the molecular mechanism of HBV integration-mediated telomerase activation. 37 It is worth noting that HBV DNA integration has been found in approximately 70% of HCC patients with occult HBV infection (OBI) in recent years, mainly involving the virus X and preS/S genomic regions. 38,39 HBV DNA gene integration is associated with changes in tumor suppressor genes, mutations in the p53 gene, and genomic instability in HCC patients with OBI.…”

Characterization of hepatitis B virus DNA integration patterns in intrahepatic cholangiocarcinoma

“…To date, 19 independent studies have identified a total of 435 integrations of HBV-DNA in the TERT gene, and 62.99% (274/435) of HBV-DNA integration occurs in the promoter region of TERT. As HBV-DNA insertion into the TERT promoter was associated with an increased TERT mRNA expression and a more aggressive tumor behavior in HCC, 36 it is hypothesized that the recurrence of HBV-DNA integrations into the TERT promoter region in HCC could be due to the potential growth advantage and clonal expansion capacity rendered by the augmented TERT expression. [37][38][39] In this study, TERT was inserted by HBV-DNA four times in three different ICC samples.…”

Hepatitis B Virus DNA Integration and Clonal Expansion of Hepatocytes in the Chronically Infected Liver

“…More recently, HCC integrations have been found in TERT, MLL4, CCNE1, and CCNA2, etc. (e.g., [35,36]), mostly in promoter/intron regions, as well as in DNA sequences for non-coding RNAs [37]. However, unlike the situation in the woodchuck model, well-defined relationships between integration and tumorigenesis have been found in less than half of human HCCs analyzed so far.…”