Role of the hepatitis B virus X protein in viral replication and transcription.

摘要

The Hepatitis B virus encodes a largely cytoplasmic, but also nuclear protein called HBx that is required for infection in vivo. HBx has been shown to activate signal transduction cascades involving Src, Ras-Raf-MAPK, Pyk2, and calcium among others. HBx also promiscuously transactivates cellular and viral transcription. In some cases, HBx activates transcription by directly interacting with bZIP transcription factor family members (e.g. ATF and CREB). Precisely how the functions of HBx converge to increase viral replication is incompletely understood.; Studies were conducted to determine the effect of HBx on HBV replication and transcription in HepG2 (hepatoblastoma) cells. The results show that at low input genome levels, HBx activates HBV replication and transcription whereas at higher input genome levels HBx minimally impacts viral transcription but still has a profound impact on HBV replication. These findings mean that HBx can activate viral replication without first activating transcription. Thus, HBx most likely exerts a post-transcriptional effect on viral replication by activating signal transduction cascades.; HBV replication, in the absence of HBx, was shown to be significantly increased when supplemental viral polymerase was co-expressed. In addition, supplemental polymerase also increased viral capsid abundance. In contrast, supplementation of WT HBV with polymerase affected neither replication nor capsid abundance. While HBx seemed to optimize polymerase function in the absence of HBx, it did not increase polymerase activity as measured by the endogenous polymerase assay.; HBx has been shown to directly interact with bZIP proteins such as CREB. Studies were aimed at investigating the role of CREB in HBV replication and transcription. When a CREB dominant negative inhibitor (A-CREB) was cotransfected with HBV, a dose dependent decrease in HBV replication was detected. Surprisingly however, a constitutively active CREB mutant (CREB_DIEDML) failed to increase HBV transcription or replication. These results suggest, but do not prove, a role for CREB in HBV transcription and replication.; Finally, HBx restricted to the nucleus retained its ability to promote viral replication, whereas cytoplasmic HBx failed to increase viral replication. Together, these results implicate CREB and the HBV polymerase respectively as transcriptional and post-transcriptional effectors of HBx-mediated activation of viral replication.