title: Intron retention may regulate expression of Epstein-Barr virus nuclear antigen 3 family genes.
abstract: The nuclear antigen 3 family genes (EBNA-3, EBNA-4, and EBNA-6) of Epstein-Barr virus (EBV) are important for EBV-induced immortalization and survival of B lymphocytes. However, little is known about how the expression of these genes is regulated. Each of the EBNA-3, EBNA-4, and EBNA-6 genes consists of two exons separated by a small intron. Reverse transcriptase PCR assays revealed that the vast majority of the EBNA-3, EBNA-4, and EBNA-6 mRNA, expressed in transfected and EBV-infected B cells, retained intron sequences. Northern blot and S1 protection assays confirmed that most of the EBNA-3 mRNA contained intron. Examination of deletion mutants of EBNA-3 indicated that the EBNA-3 protein was not necessary for intron retention and that there was no splicing silencing element encoded in the EBNA-3 mRNA. Cell fractionation and RNA gradient analysis revealed that the unspliced EBNA 3 family mRNAs were transported into the cytoplasm and associated with the polysomes. However, Western blot analysis of FLAG-epitope tagged EBNA-3 gave no indication of the presence of splice variant protein forms of EBNA-3. In contrast, transiently transfected cells expressing EBNA-3 revealed a sixfold increase in EBNA-3 protein expression from the genomic EBNA-3 gene compared to EBNA-3 cDNA. These data show that the intronic sequences can influence EBNA-3 protein expression and suggest that intron retention may provide a means for the fine-tuning of expression of the individual EBNA 3 family genes.
authors.affiliation: EBV Unit, The Queensland Institute of Medical Research and University of Queensland Joint Oncology Program, Brisbane, Australia. norbertK@qimr.edu.au
authors.forename: N
authors.initials: N
authors.lastname: Kienzle
authors.forename: D B
authors.initials: DB
authors.lastname: Young
authors.forename: D
authors.initials: D
authors.lastname: Liaskou
authors.forename: M
authors.initials: M
authors.lastname: Buck
authors.forename: S
authors.initials: S
authors.lastname: Greco
authors.forename: T B
authors.initials: TB
authors.lastname: Sculley
mesh_terms: D017398:Alternative Splicing; D001483:Base Sequence; D001692:Biological Transport; D015152:Blotting, Northern; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism; D019309:Epstein-Barr Virus Nuclear Antigens / Q000235:genetics*; D015967:Gene Expression Regulation, Viral; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D007438:Introns; D008969:Molecular Sequence Data; D011132:Polyribosomes; D012326:RNA Splicing; D012333:RNA, Messenger; D012367:RNA, Viral; D015719:Single-Strand Specific DNA and RNA Endonucleases
chemical_list: D019309:Epstein-Barr Virus Nuclear Antigens; D012333:RNA, Messenger; D012367:RNA, Viral; D015719:Single-Strand Specific DNA and RNA Endonucleases