Characterization of the transcriptional repressor RBP in Epstein-Barr virus-transformed B cells.
RBP, a transcriptional repressor, is intricately involved in Epstein-Barr virus (EBV) transformation of human B cells. The EBV nuclear proteins EBNA-2, -3, -4 and -6 all utilize RBP to regulate the transcription of both cellular and viral genes. This study investigates the isoforms of the RBP protein in Burkitt's lymphoma (BL) cells and in EBV-transformed lymphoblastoid cell lines (LCLs). Two-dimensional gel electrophoresis showed the presence of two different cellular isoforms of RBP; the molecular masses and isoelectric points of these two isoforms corresponded to RBP-Jkappa and RBP-2N. Fractionation studies and green fluorescent protein (GFP)-tagged expression studies demonstrated that both RBP isoforms were located predominantly in the cell nucleus. Interestingly, GFP-tagged RBP-Jkappa showed diffuse, uniform nuclear staining, whereas GFP-tagged RBP-2N showed a discrete nuclear pattern, demonstrating differences between the two isoforms. Within the nuclear fraction of EBV-negative BL cells, RBP existed both in a free form and bound to chromatin, whereas in LCLs the intranuclear RBP was predominantly chromatin-bound. Expression of the EBV latent proteins was found to lead to the sequestering of RBP from the cytoplasm into the cell nucleus and to an increase in the chromatin-bound forms of RBP.
Krauer
Kenia G
KG
Queensland Institute of Medical Research and University of Queensland Joint Oncology Program, 300 Herston Road, Herston 4029, Brisbane, Australia1.
Buck
Marion
M
Queensland Institute of Medical Research and University of Queensland Joint Oncology Program, 300 Herston Road, Herston 4029, Brisbane, Australia1.
Sculley
Tom
T
Queensland Institute of Medical Research and University of Queensland Joint Oncology Program, 300 Herston Road, Herston 4029, Brisbane, Australia1.
D001402:B-Lymphocytes / Q000378:metabolism / Q000821:virology*; D002461:Cell Line, Transformed; D002467:Cell Nucleus / Q000378:metabolism; D002472:Cell Transformation, Viral; D002843:Chromatin / Q000378:metabolism; D016384:Consensus Sequence; D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D019309:Epstein-Barr Virus Nuclear Antigens / Q000378:metabolism; D015967:Gene Expression Regulation, Viral; D049452:Green Fluorescent Proteins; D004854:Herpesvirus 4, Human / Q000235:genetics* / Q000378:metabolism; D008164:Luminescent Proteins / Q000378:metabolism; D009687:Nuclear Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D017735:Virus Latency / Q000235:genetics
D002843:Chromatin; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D008164:Luminescent Proteins; D009687:Nuclear Proteins; D020033:Protein Isoforms; D011993:Recombinant Fusion Proteins; D012097:Repressor Proteins; D049452:Green Fluorescent Proteins; D004247:DNA