LAUSR

Hepatitis B virus (HBV) integrations are common in hepatocellular carcinoma (HCC). In particular, alterations of TERT gene by HBV integrations are frequent; however the molecular mechanism and functional consequence underlying TERT-HBV integration are unclear. 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 upon 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 siRNA 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 upon viral integration. We identified a previously unrecognized molecular mechanism of TERT activation via 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 novel 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.