Epstein-Barr virus encoded nuclear protein EBNA-3 binds a novel human uridine kinase/uracil phosphoribosyltransferase
Epstein-Barr virus encoded nuclear protein EBNA-3 binds a novel human uridine kinase/uracil phosphoribosyltransferase
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Abstract Background Epstein-Barr virus (EBV) infects resting B-lymphocytes and transforms them OTF into immortal proliferating lymphoblastoid cell lines (LCLs) in vitro.The transformed immunoblasts may grow up as immunoblastic lymphomas in immuno-suppressed hosts.Results In order to identify cellular protein targets that may be involved in Epstein-Barr virus mediated B-cell transformation, human LCL cDNA library was screened with one of the transformation associated nuclear antigens, EBNA-3 (also called EBNA-3A), using the yeast two-hybrid system.
A clone encoding a fragment of a novel human protein was isolated (clone 538).The interaction was confirmed using in vitro binding assays.A full-length cDNA clone (F538) was isolated.
Sequence alignment with known proteins and 3D structure predictions suggest that F538 is a novel human uridine kinase/uracil phosphoribosyltransferase.The GFP-F538 fluorescent fusion protein showed a preferentially cytoplasmic distribution but translocated to the nucleus upon co-expression of EBNA-3.A naturally occurring splice variant of F538, that lacks the C-terminal uracil phosphoribosyltransferase Waxed Cotton Dressing part but maintain uridine kinase domain, did not translocate to the nucleus in the presence of EBNA3.
Antibody that was raised against the bacterially produced GST-538 protein showed cytoplasmic staining in EBV negative Burkitt lymphomas but gave a predominantly nuclear staining in EBV positive LCL-s and stable transfected cells expressing EBNA-3.Conclusion We suggest that EBNA-3 by direct protein-potein interaction induces the nuclear accumulation of a novel enzyme, that is part of the ribonucleotide salvage pathway.Increased intranuclear levels of UK/UPRT may contribute to the metabolic build-up that is needed for blast transformation and rapid proliferation.