The hepatitis B virus HBx protein is a dual specificity cytoplasmic activator of Ras and nuclear activator of transcription factors.
The HBx protein of hepatitis B virus (HBV) is a transcriptional activator that is required for infection and may play an important role in HBV-associated hepatocarcinogenesis. Recently, we and others have shown that HBx stimulates the Ras-Raf-MAP kinase cascade, which leads to enhanced cell proliferation and the activation of transcription factors AP-1 and NF-kappa B. Other studies have shown that HBx can activate transcription by interacting directly with nuclear components of the transcription machinery. Therefore we examined the basis for the different reported activities of HBx. Here, we show that HBx is a complex protein, displaying independent activities in different intracellular locations. The intracellular distribution of HBx protein was first investigated using scanning confocal laser immunomicroscopy and by genetic studies. Our work has established that HBx expressed in cultured cells is found authentically in both the cytoplasm and the nucleus. HBx is not strongly associated with any intracellular structures, but some preferential accumulation was observed near the cell surface. Next, HBx variants were constructed containing a functional or mutant nuclear localization sequence. We show that when HBx is engineered to relocate exclusively to the nucleus, it no longer activates the Ras-Raf-MAP kinase cascade, nor does it activate transcription factors AP-1 and NF-kappa B. Surprisingly, nuclear HBx fully retains the ability to stimulate HBV enhancer I, which is activated independently of the Ras and protein kinase C pathways. Therefore HBx protein stimulates signal transduction pathways in the cytoplasm and transactivates transcription elements in the nucleus. Furthermore, SV40 T antigen is shown to induce the nuclear sequestration of HBx protein and to block its activation of NF-kappa B, demonstrating that HBx is regulated by proteins that alter its intracellular distribution. The conflicting functions of HBx protein in viral infection and possibly carcinoma may involve the regulation of its differential distribution in the cell.
Doria
M
M
Department of Biochemistry, New York University School of Medicine, NY 10016, USA.
Klein
N
N
Lucito
R
R
Schneider
R J
RJ
D000595:Amino Acid Sequence; D000952:Antigens, Polyomavirus Transforming / Q000502:physiology; D001483:Base Sequence; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000502:physiology; D002467:Cell Nucleus / Q000737:chemistry*; D003593:Cytoplasm / Q000737:chemistry*; D004742:Enhancer Elements, Genetic; D006515:Hepatitis B virus; D006801:Humans; D008969:Molecular Sequence Data; D016328:NF-kappa B; D017346:Protein Serine-Threonine Kinases / Q000502:physiology; D011518:Proto-Oncogene Proteins / Q000502:physiology; D019908:Proto-Oncogene Proteins c-raf; D016283:Proto-Oncogene Proteins p21(ras); D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D015398:Signal Transduction / Q000502:physiology*; D013539:Simian virus 40 / Q000276:immunology; D015534:Trans-Activators / Q000032:analysis* / Q000235:genetics / Q000502:physiology; D018808:Transcription Factor AP-1; D015533:Transcriptional Activation / Q000502:physiology; D054334:Viral Regulatory and Accessory Proteins
D000952:Antigens, Polyomavirus Transforming; D016328:NF-kappa B; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D018808:Transcription Factor AP-1; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C501009:HRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)