epstein-barr virus protocols

Methods in Molecular Biology TM HUMANA PRESS Epstein-Barr Virus Protocols Edited by Joanna B. Wilson Gerhard H. W. May HUMANA PRESS Methods in Molecular Biology TM VOLUME 174 Epstein-Barr Virus Protocols Edited by Joanna B. Wilson Gerhard H. W. May EBV: B95-8 Strain Map 3 3 From: Methods in Molecular Biology, Vol. 174: Epstein-Barr Virus Protocols Edited by: J. B. Wilson and G. H. W. May © Humana Press Inc., Totowa, NJ 1 Epstein-Barr Virus The B95-8 Strain Map Paul J. Farrell 1. Introduction This chapter summarizes the genes and mRNAs that have been mapped on to the B95-8 EBV genome. The complete sequence of this strain of Epstein-Barr Virus (EBV) was established (1,2) and data from many publications has been integrated into the map, which is an update of that published previously (3). The B95-8 strain grows well in laboratory culture and transforms human B lymphocytes efficiently, but B95-8 EBV has about 11.8 kb deleted relative to other strains of EBV. The sequence of that region has been determined in the Raji strain of EBV and a map has been published (4). Detailed literature citations for most of the features have been published (3). The map should be used in conjunction with the B95-8 EBV DNA sequence, which can be accessed in the European Molecular Biology Laboratory (EMBL; embl:ebv.seq) or Genbank databases. The feature tables shown in those database files have not yet been revised so the information shown in this map is considerably more up to date. 2. Repeat Sequences There are several regions of the genome that contain tandem repeat sequences. Some of these are large repeat units (for example, the major internal repeat is 3072 bp and the terminal repeat is 538 bp) but some are much simpler repeats. In most virus preparations, there will be a distribution of copy number of these repeats, so it is impor- tant to appreciate that the viral map and coordinates are just one reference example of genome structure. Generally the repeat numbers in the B95-8 map are thought to be typical, although it now appears that the 11.5 copies of the major internal repeat inserted into the B95-8 sequence may be an over-estimate, about 8.5 being more usual in B95-8 EBV (5). The restriction maps for EcoRI and BamHI are shown beneath the scale bar in kb. The restriction fragments are labeled according to size, A being the largest fragment 4Farrell and lower case letters being used for additional fragments when there are more than 26. 3. Open Reading Frames The major open reading frames, presumed to correspond to the protein coding parts of genes are shown as boxes, pointed at one end to indicate the direction of translation. Coordinates shown below are generally from the presumed initiator methionine to the last translated amino acid. When there is no obvious initiator methionine or the RNA structure is uncertain, the coordinates of the reading frame are shown from the begin- ning of the open reading frame to the last translated amino acid; these reading frames are marked “orf” after the coordinates. Initiator methionine residues have generally not been confirmed by mutagenesis so it is important to appreciate that the map is an interpretation in this respect. 4. Nomenclature Open reading frames of EBV are named systematically according to the BamHI restriction fragment in which their transcription commences, designated L or R accord- ing to the direction and then marked F followed by a number. So BALF4 is the fourth leftward reading frame commencing in the BamHI A restriction fragment. Many of the genes are also known by more common and useful names, which may describe their function. Abbreviations used here are EBNA, Epstein-Barr virus nuclear antigen, LMP, latent membrane protein, EBER, Epstein-Barr virus encoded RNA. There is unfortunately some variation in nomenclature for EBV genes in the published litera- ture. A guide to equivalents is shown in Table 1; terms on the same horizontal line refer to the same gene that shown on the left, being the standard used in this map. 5. Gene Expression The open reading frames are shaded according to their expression class. These classes are defined operationally in EBV. Latent cycle genes are expressed constitu- Table 1 EBV Gene Nomenclature EBNA-3A EBNA-3 EBNA-3B EBNA-4 EBNA-3C EBNA-6 EBNA-LP EBNA-5 LMP-2A TP-1 LMP-2B TP-2 BZLF1 ZEBRA, EB1, Zta BRLF1 Rta, R BARF transcripts BART, CST EBNA, Epstein-Barr virus nuclear antigen; LMP, latent membrane protein; TP, terminal protein; BART, Bam A rightward transcripts; CST, complementary strand transcripts. (text continued on p. 12) EBV: B95-8 Strain Map 5 6Farrell EBV: B95-8 Strain Map 7 8Farrell EBV: B95-8 Strain Map 9 10 Farrell EBV: B95-8 Strain Map 11 [...]... Dominant-negative inhibitors of EBNA-1 of Epstein-Barr virus J Virol 71, 1766–1775 2 Kirchmaier, A L and Sugden, B (1998) Rep*: a viral element that can partially replace the origin of plasmid DNA synthesis of EBV J Virol 72, 4657–4666 3 Lupton, S and Levine, A J (1985) Mapping genetic elements of Epstein-Barr virus that facilitate extrachromosomal persistence of Epstein-Barr virus- derived plasmids in human... Raji Epstein-Barr virus DNA spanning the B95-8 deletion Virology 179, 339–346 5 Allan, G J and Rowe, D T (1989) Size and stability of the Epstein-Barr virus major internal repeat (IR-1) in Burkitt’s lymphoma and lymphoblastoid cell lines Virology 173, 489–498 6 Ganem, D (1997) KSHV and Kaposi’s Sarcoma: the end of the beginning? Cell 91, 157–160 7 McGeoch, D J and Schaffer, P A (1993) Herpes Simplex Virus, ... plasmids derived from Epstein-Barr virus is composed of two cis-acting components Mol Cell Biol 5, 1822–1832 5 Sugden, B., Marsh, K., and Yates, J (1985) A vector that replicates as a plasmid and can be efficiently selected in B-lymphocytes transformed by Epstein-Barr virus Mol Cell Biol 5, 410–413 6 Yates, J., Warren, N., Reisman, D., and Sugden, B (1984) A cis-acting element from the Epstein-Barr viral... irradiated feeder cells can be immediately mixed with primary B-cells and virus- stock References 1 Cohen, J I., Wang, F., Mannick, J., and Kieff, E (1989) Epstein-Barr virus nuclear protein 2 is a key determinant of lymphocyte transformation Proc Natl Acad Sci USA 86, 9558–9562 2 Kaye, K M., Izumi, K M., and Kieff, E (1993) Epstein-Barr virus latent membrane protein 1 is essential for B-lymphocyte growth... B-lymphocytes by a plasmid containing 71 kpb of Epstein-Barr viral DNA J Virol 69, 231–238 9 Rabson, M., Gradoville, L., Heston, L., and Miller, G (1982) Non-immortalizing P3JHR-1 Epstein-Barr virus: a deletion mutant of its transforming parent, Jijoye J Virol 44, 834–844 10 Kilger, E., Kieser, A., Baumann, M., and Hammerschmidt, W (1998) Epstein-Barr virusmediated B-cell proliferation is dependent... 109–135 16 Heston, L., Rabson, M., Brown, N., and Miller, G (1982) New Epstein-Barr virus variants from cellular subclones of P3J-HR-1 Burkitt lymphoma Nature 295, 160–163 Mini-EBV Virus Plasmids 35 17 Parker, B D., Bankier, A., Satchwell, S., Barrell, B., and Farrell, P J (1990) Sequence and transcription of Raji Epstein-Barr virus DNA spanning the B95-8 deletion region Virology 179, 339–346 18 Southern,... activates transcription but not replication J Virol 66, 1795–1798 Mini-EBV Virus Plasmids 23 3 Genetic Analysis and Gene Expression with Mini -Epstein-Barr Virus Plasmids Ellen Kilger and Wolfgang Hammerschmidt 1 Introduction Upon infection with Epstein-Barr virus (EBV), primary human B-lymphocytes are efficiently immortalized and give rise to lymphoblastoid cell lines in vitro Four of the 11 viral... plasmid can then be packaged into virions in the helper cell line HH514, which carries the endogenous nonimmortalizing EBV strain P3HR1 (9) These virus- stocks can be used to immortalize primary B-cells From: Methods in Molecular Biology, Vol 174: Epstein-Barr Virus Protocols Edited by: J B Wilson and G H W May © Humana Press Inc., Totowa, NJ 23 24 Kilger and Hammerschmidt Fig 1 Composition of the mini-EBV... Gibson, T J., et al (1984) DNA sequence and expression of the B95–8 Epstein-Barr virus genome Nature 310, 207–211 2 Laux, G., Perricaudet, M., and Farrell, P J (1988) A spliced EBV gene expressed in immortalised lymphocytes is created by circularisation of the linear viral genome EMBO J 7, 769–774 3 Farrell P J (1993) Epstein-Barr Virus, in Genetic Maps, 6th ed (O’Brien, S J., ed.), Cold Spring Harbor... Proc Natl Acad Sci USA 90, 9150–9154 3 Tomkinson, B., Robertson, E., and Kieff, E (1993) Epstein-Barr virus nuclear proteins EBNA-3A and EBNA-3C are essential for B-lymphocyte growth transformation J Virol 67, 2014–2025 4 Hammerschmidt, W and Sugden, B (1989) Genetic analysis of immortalizing functions of Epstein-Barr virus in human B lymphocytes Nature 340, 393–397 5 Kempkes, B., Spitkovsky, D., Jansen-Dürr, . Biology TM HUMANA PRESS Epstein-Barr Virus Protocols Edited by Joanna B. Wilson Gerhard H. W. May HUMANA PRESS Methods in Molecular Biology TM VOLUME 174 Epstein-Barr Virus Protocols Edited by Joanna. 3 3 From: Methods in Molecular Biology, Vol. 174: Epstein-Barr Virus Protocols Edited by: J. B. Wilson and G. H. W. May © Humana Press Inc., Totowa, NJ 1 Epstein-Barr Virus The B95-8 Strain Map Paul J. Farrell 1 describe their function. Abbreviations used here are EBNA, Epstein-Barr virus nuclear antigen, LMP, latent membrane protein, EBER, Epstein-Barr virus encoded RNA. There is unfortunately some variation