Epstein-Barr virus genetics: talking about the BAC generation

Herpesviridae - Tập 1 - Trang 1-13 - 2010
Regina Feederle1, Emmalene J Bartlett1, Henri-Jacques Delecluse1
1German Cancer Research Centre, Heidelberg, Germany

Tóm tắt

Genetic mutant organisms pervade all areas of Biology. Early on, herpesviruses (HV) were found to be amenable to genetic analysis using homologous recombination techniques in eukaryotic cells. More recently, HV genomes cloned onto a bacterial artificial chromosome (BAC) have become available. HV BACs can be easily modified in E.coli and reintroduced in eukaryotic cells to produce infectious viruses. Mutants derived from HV BACs have been used both to understand the functions of all types of genetic elements present on the virus genome, but also to generate mutants with potentially medically relevant properties such as preventative vaccines. Here we retrace the development of the BAC technology applied to the Epstein-Barr virus (EBV) and review the strategies available for the construction of mutants. We expand on the appropriate controls required for proper use of the EBV BACs, and on the technical hurdles researchers face in working with these recombinants. We then discuss how further technological developments might successfully overcome these difficulties. Finally, we catalog the EBV BAC mutants that are currently available and illustrate their contributions to the field using a few representative examples.

Tài liệu tham khảo

Heller M, Dambaugh T, Kieff E: Epstein-Barr virus DNA. IX. Variation among viral DNAs from producer and nonproducer infected cells. J Virol. 1981, 38: 632-648. Polack A, Delius H, Zimber U, Bornkamm GW: Two deletions in the Epstein-Barr virus genome of the Burkitt lymphoma nonproducer line Raji. Virology. 1984, 133: 146-157. 10.1016/0042-6822(84)90433-1. Rabson M, Gradoville L, Heston L, Miller G: Non-immortalizing P3J-HR-1 Epstein-Barr virus: a deletion mutant of its transforming parent, Jijoye. J Virol. 1982, 44: 834-844. Hammerschmidt W, Sugden B: Genetic analysis of immortalizing functions of Epstein-Barr virus in human B lymphocytes. Nature. 1989, 340: 393-397. 10.1038/340393a0. Cohen JI, Wang F, Mannick J, Kieff E: Epstein-Barr virus nuclear protein 2 is a key determinant of lymphocyte transformation. Proc Natl Acad Sci USA. 1989, 86: 9558-9562. 10.1073/pnas.86.23.9558. Tomkinson B, Kieff E: Use of second-site homologous recombination to demonstrate that Epstein-Barr virus nuclear protein 3B is not important for lymphocyte infection or growth transformation in vitro. J Virol. 1992, 66: 2893-2903. Shimizu N, Yoshiyama H, Takada K: Clonal propagation of Epstein-Barr virus (EBV) recombinants in EBV-negative Akata cells. J Virol. 1996, 70: 7260-7263. O'Connor M, Peifer M, Bender W: Construction of large DNA segments in Escherichia coli. Science. 1989, 244: 1307-1312. Messerle M, Crnkovic I, Hammerschmidt W, Ziegler H, Koszinowski UH: Cloning and mutagenesis of a herpesvirus genome as an infectious bacterial artificial chromosome. Proc Natl Acad Sci USA. 1997, 94: 14759-14763. 10.1073/pnas.94.26.14759. Delecluse HJ, Hilsendegen T, Pich D, Zeidler R, Hammerschmidt W: Propagation and recovery of intact, infectious Epstein-Barr virus from prokaryotic to human cells. Proc Natl Acad Sci USA. 1998, 95: 8245-8250. 10.1073/pnas.95.14.8245. Borst EM, Hahn G, Koszinowski UH, Messerle M: Cloning of the human cytomegalovirus (HCMV) genome as an infectious bacterial artificial chromosome in Escherichia coli: a new approach for construction of HCMV mutants. J Virol. 1999, 73: 8320-8329. Adler H, Messerle M, Wagner M, Koszinowski UH: Cloning and mutagenesis of the murine gammaherpesvirus 68 genome as an infectious bacterial artificial chromosome. J Virol. 2000, 74: 6964-6974. 10.1128/JVI.74.15.6964-6974.2000. Saeki Y, Ichikawa T, Saeki A, Chiocca EA, Tobler K, Ackermann M, Breakefield XO, Fraefel C: Herpes simplex virus type 1 DNA amplified as bacterial artificial chromosome in Escherichia coli: rescue of replication-competent virus progeny and packaging of amplicon vectors. Hum Gene Ther. 1998, 9: 2787-2794. 10.1089/hum.1998.9.18-2787. Nagaike K, Mori Y, Gomi Y, Yoshii H, Takahashi M, Wagner M, Koszinowski U, Yamanishi K: Cloning of the varicella-zoster virus genome as an infectious bacterial artificial chromosome in Escherichia coli. Vaccine. 2004, 22: 4069-4074. 10.1016/j.vaccine.2004.03.062. Delecluse HJ, Kost M, Feederle R, Wilson L, Hammerschmidt W: Spontaneous activation of the lytic cycle in cells infected with a recombinant Kaposi's sarcoma-associated virus. J Virol. 2001, 75: 2921-2928. 10.1128/JVI.75.6.2921-2928.2001. Zhou FC, Zhang YJ, Deng JH, Wang XP, Pan HY, Hettler E, Gao SJ: Efficient infection by a recombinant Kaposi's sarcoma-associated herpesvirus cloned in a bacterial artificial chromosome: application for genetic analysis. J Virol. 2002, 76: 6185-6196. 10.1128/JVI.76.12.6185-6196.2002. Chang WL, Barry PA: Cloning of the full-length rhesus cytomegalovirus genome as an infectious and self-excisable bacterial artificial chromosome for analysis of viral pathogenesis. J Virol. 2003, 77: 5073-5083. 10.1128/JVI.77.9.5073-5083.2003. Estep RD, Powers MF, Yen BK, Li H, Wong SW: Construction of an infectious rhesus rhadinovirus bacterial artificial chromosome for the analysis of Kaposi's sarcoma-associated herpesvirus-related disease development. J Virol. 2007, 81: 2957-2969. 10.1128/JVI.01997-06. Smith GA, Enquist LW: Construction and transposon mutagenesis in Escherichia coli of a full-length infectious clone of pseudorabies virus, an alphaherpesvirus. J Virol. 1999, 73: 6405-6414. White RE, Calderwood MA, Whitehouse A: Generation and precise modification of a herpesvirus saimiri bacterial artificial chromosome demonstrates that the terminal repeats are required for both virus production and episomal persistence. J Gen Virol. 2003, 84: 3393-3403. 10.1099/vir.0.19387-0. Petherbridge L, Brown AC, Baigent SJ, Howes K, Sacco MA, Osterrieder N, Nair VK: Oncogenicity of virulent Marek's disease virus cloned as bacterial artificial chromosomes. J Virol. 2004, 78: 13376-13380. 10.1128/JVI.78.23.13376-13380.2004. Kanda T, Yajima M, Ahsan N, Tanaka M, Takada K: Production of high-titer Epstein-Barr virus recombinants derived from Akata cells by using a bacterial artificial chromosome system. J Virol. 2004, 78: 7004-7015. 10.1128/JVI.78.13.7004-7015.2004. Chen A, Divisconte M, Jiang X, Quink C, Wang F: Epstein-Barr virus with the latent infection nuclear antigen 3B completely deleted is still competent for B-cell growth transformation in vitro. J Virol. 2005, 79: 4506-4509. 10.1128/JVI.79.7.4506-4509.2005. Izumi KM: Identification of EBV transforming genes by recombinant EBV technology. Semin Cancer Biol. 2001, 11: 407-414. 10.1006/scbi.2001.0407. Kieff ED, Rickinson AB: Epstein-Barr Virus and its replication. Fields Virology. Edited by: Knipe DM HP, Griffin DE, Lamb RA, Martin MA, Roizman B, Straus SE. 2007, Philadelphia: Lippincott Williams & Wilkins, 2: 2603-2654. 5 Horsburgh BC, Hubinette MM, Qiang D, MacDonald ML, Tufaro F: Allele replacement: an application that permits rapid manipulation of herpes simplex virus type 1 genomes. Gene Ther. 1999, 6: 922-930. 10.1038/sj.gt.3300887. Feng WH, Cohen JI, Fischer S, Li L, Sneller M, Goldbach-Mansky R, Raab-Traub N, Delecluse HJ, Kenney SC: Reactivation of latent Epstein-Barr virus by methotrexate: a potential contributor to methotrexate-associated lymphomas. J Natl Cancer Inst. 2004, 96: 1691-1702. 10.1093/jnci/djh313. Takada K: Cross-linking of cell surface immunoglobulins induces Epstein-Barr virus in Burkitt lymphoma lines. Int J Cancer. 1984, 33: 27-32. 10.1002/ijc.2910330106. Neuhierl B, Delecluse HJ: Molecular genetics of DNA viruses: recombinant virus technology. Methods Mol Biol. 2005, 292: 353-370. Poteete AR: What makes the bacteriophage lambda Red system useful for genetic engineering: molecular mechanism and biological function. FEMS Microbiol Lett. 2001, 201: 9-14. Zhang Y, Buchholz F, Muyrers JP, Stewart AF: A new logic for DNA engineering using recombination in Escherichia coli. Nat Genet. 1998, 20: 123-128. 10.1038/2417. Warming S, Costantino N, Court DL, Jenkins NA, Copeland NG: Simple and highly efficient BAC recombineering using galK selection. Nucleic Acids Res. 2005, 33: e36-10.1093/nar/gni035. Wang S, Zhao Y, Leiby M, Zhu J: A new positive/negative selection scheme for precise BAC recombineering. Mol Biotechnol. 2009, 42: 110-116. 10.1007/s12033-009-9142-3. Tischer BK, von Einem J, Kaufer B, Osterrieder N: Two-step red-mediated recombination for versatile high-efficiency markerless DNA manipulation in Escherichia coli. Biotechniques. 2006, 40: 191-197. 10.2144/000112096. Brune W, Menard C, Hobom U, Odenbreit S, Messerle M, Koszinowski UH: Rapid identification of essential and nonessential herpesvirus genes by direct transposon mutagenesis. Nat Biotechnol. 1999, 17: 360-364. 10.1038/7914. Le Clorennec C, Ouk TS, Youlyouz-Marfak I, Panteix S, Martin CC, Rastelli J, Adriaenssens E, Zimber-Strobl U, Coll J, Feuillard J, et al: Molecular basis of cytotoxicity of Epstein-Barr virus (EBV) latent membrane protein 1 (LMP1) in EBV latency III B cells: LMP1 induces type II ligand-independent autoactivation of CD95/Fas with caspase 8-mediated apoptosis. J Virol. 2008, 82: 6721-6733. 10.1128/JVI.02250-07. Croft NP, Shannon-Lowe C, Bell AI, Horst D, Kremmer E, Ressing ME, Wiertz EJ, Middeldorp JM, Rowe M, Rickinson AB, et al: Stage-specific inhibition of MHC class I presentation by the Epstein-Barr virus BNLF2a protein during virus lytic cycle. PLoS Pathog. 2009, 5: e1000490-10.1371/journal.ppat.1000490. Feederle R, Mehl-Lautscham AM, Bannert H, Delecluse HJ: The Epstein-Barr virus protein kinase BGLF4 and the exonuclease BGLF5 have opposite effects on the regulation of viral protein production. J Virol. 2009, 83: 10877-10891. 10.1128/JVI.00525-09. Bornkamm GW, Berens C, Kuklik-Roos C, Bechet JM, Laux G, Bachl J, Korndoerfer M, Schlee M, Holzel M, Malamoussi A, et al: Stringent doxycycline-dependent control of gene activities using an episomal one-vector system. Nucleic Acids Res. 2005, 33: e137-10.1093/nar/gni137. Kintner C, Sugden B: Conservation and progressive methylation of Epstein-Barr viral DNA sequences in transformed cells. J Virol. 1981, 38: 305-316. Kalla M, Schmeinck A, Bergbauer M, Pich D, Hammerschmidt W: AP-1 homolog BZLF1 of Epstein-Barr virus has two essential functions dependent on the epigenetic state of the viral genome. Proc Natl Acad Sci USA. 107: 850-855. 10.1073/pnas.0911948107. Janz A, Oezel M, Kurzeder C, Mautner J, Pich D, Kost M, Hammerschmidt W, Delecluse HJ: Infectious Epstein-Barr virus lacking major glycoprotein BLLF1 (gp350/220) demonstrates the existence of additional viral ligands. J Virol. 2000, 74: 10142-10152. 10.1128/JVI.74.21.10142-10152.2000. Neuhierl B, Feederle R, Adhikary D, Hub B, Geletneky K, Mautner J, Delecluse HJ: Primary B-cell infection with a deltaBALF4 Epstein-Barr virus comes to a halt in the endosomal compartment yet still elicits a potent CD4-positive cytotoxic T-cell response. J Virol. 2009, 83: 4616-4623. 10.1128/JVI.01613-08. Busse C, Feederle R, Schnolzer M, Behrends U, Mautner J, Delecluse HJ: Epstein-Barr viruses that express a CD21 antibody provide evidence that gp350's functions extend beyond B-cell surface binding. J Virol. 2010, 84: 1139-1147. 10.1128/JVI.01953-09. Countryman J, Miller G: Activation of expression of latent Epstein-Barr herpesvirus after gene transfer with a small cloned subfragment of heterogeneous viral DNA. Proc Natl Acad Sci USA. 1985, 82: 4085-4089. 10.1073/pnas.82.12.4085. Hardwick JM, Lieberman PM, Hayward SD: A new Epstein-Barr virus transactivator, R, induces expression of a cytoplasmic early antigen. J Virol. 1988, 62: 2274-2284. Feederle R, Kost M, Baumann M, Janz A, Drouet E, Hammerschmidt W, Delecluse HJ: The Epstein-Barr virus lytic program is controlled by the co-operative functions of two transactivators. Embo J. 2000, 19: 3080-3089. 10.1093/emboj/19.12.3080. Deng Z, Chen CJ, Zerby D, Delecluse HJ, Lieberman PM: Identification of acidic and aromatic residues in the Zta activation domain essential for Epstein-Barr virus reactivation. J Virol. 2001, 75: 10334-10347. 10.1128/JVI.75.21.10334-10347.2001. Bhende PM, Seaman WT, Delecluse HJ, Kenney SC: The EBV lytic switch protein, Z, preferentially binds to and activates the methylated viral genome. Nat Genet. 2004, 36: 1099-1104. 10.1038/ng1424. Bhende PM, Seaman WT, Delecluse HJ, Kenney SC: BZLF1 activation of the methylated form of the BRLF1 immediate-early promoter is regulated by BZLF1 residue 186. J Virol. 2005, 79: 7338-7348. 10.1128/JVI.79.12.7338-7348.2005. Schelcher C, Valencia S, Delecluse HJ, Hicks M, Sinclair AJ: Mutation of a single amino acid residue in the basic region of the Epstein-Barr virus (EBV) lytic cycle switch protein Zta (BZLF1) prevents reactivation of EBV from latency. J Virol. 2005, 79: 13822-13828. 10.1128/JVI.79.21.13822-13828.2005. Heston L, El-Guindy A, Countryman J, Dela Cruz C, Delecluse HJ, Miller G: Amino acids in the basic domain of Epstein-Barr virus ZEBRA protein play distinct roles in DNA binding, activation of early lytic gene expression, and promotion of viral DNA replication. J Virol. 2006, 80: 9115-9133. 10.1128/JVI.00909-06. El-Guindy A, Heston L, Delecluse HJ, Miller G: Phosphoacceptor site S173 in the regulatory domain of Epstein-Barr Virus ZEBRA protein is required for lytic DNA replication but not for activation of viral early genes. J Virol. 2007, 81: 3303-3316. 10.1128/JVI.02445-06. Hong GK, Kumar P, Wang L, Damania B, Gulley ML, Delecluse HJ, Polverini PJ, Kenney SC: Epstein-Barr virus lytic infection is required for efficient production of the angiogenesis factor vascular endothelial growth factor in lymphoblastoid cell lines. J Virol. 2005, 79: 13984-13992. 10.1128/JVI.79.22.13984-13992.2005. Park R, Heston L, Shedd D, Delecluse HJ, Miller G: Mutations of amino acids in the DNA-recognition domain of Epstein-Barr virus ZEBRA protein alter its sub-nuclear localization and affect formation of replication compartments. Virology. 2008, 382: 145-162. 10.1016/j.virol.2008.09.009. Jones RJ, Seaman WT, Feng WH, Barlow E, Dickerson S, Delecluse HJ, Kenney SC: Roles of lytic viral infection and IL-6 in early versus late passage lymphoblastoid cell lines and EBV-associated lymphoproliferative disease. Int J Cancer. 2007, 121: 1274-1281. 10.1002/ijc.22839. Pudney VA, Leese AM, Rickinson AB, Hislop AD: CD8+ immunodominance among Epstein-Barr virus lytic cycle antigens directly reflects the efficiency of antigen presentation in lytically infected cells. J Exp Med. 2005, 201: 349-360. 10.1084/jem.20041542. Zuo J, Currin A, Griffin BD, Shannon-Lowe C, Thomas WA, Ressing ME, Wiertz EJ, Rowe M: The Epstein-Barr virus G-protein-coupled receptor contributes to immune evasion by targeting MHC class I molecules for degradation. PLoS Pathog. 2009, 5: e1000255-10.1371/journal.ppat.1000255. Humme S, Reisbach G, Feederle R, Delecluse HJ, Bousset K, Hammerschmidt W, Schepers A: The EBV nuclear antigen 1 (EBNA1) enhances B cell immortalization several thousandfold. Proc Natl Acad Sci USA. 2003, 100: 10989-10994. 10.1073/pnas.1832776100. Altmann M, Pich D, Ruiss R, Wang J, Sugden B, Hammerschmidt W: Transcriptional activation by EBV nuclear antigen 1 is essential for the expression of EBV's transforming genes. Proc Natl Acad Sci USA. 2006, 103: 14188-14193. 10.1073/pnas.0605985103. Altmann M, Hammerschmidt W: Epstein-Barr virus provides a new paradigm: a requirement for the immediate inhibition of apoptosis. PLoS Biol. 2005, 3: e404-10.1371/journal.pbio.0030404. Rowe M, Glaunsinger B, van Leeuwen D, Zuo J, Sweetman D, Ganem D, Middeldorp J, Wiertz EJ, Ressing ME: Host shutoff during productive Epstein-Barr virus infection is mediated by BGLF5 and may contribute to immune evasion. Proc Natl Acad Sci USA. 2007, 104: 3366-3371. 10.1073/pnas.0611128104. Zuo J, Thomas W, van Leeuwen D, Middeldorp JM, Wiertz EJ, Ressing ME, Rowe M: The DNase of gammaherpesviruses impairs recognition by virus-specific CD8+ T cells through an additional host shutoff function. J Virol. 2008, 82: 2385-2393. 10.1128/JVI.01946-07. Ressing ME, van Leeuwen D, Verreck FA, Keating S, Gomez R, Franken KL, Ottenhoff TH, Spriggs M, Schumacher TN, Hutt-Fletcher LM, et al: Epstein-Barr virus gp42 is posttranslationally modified to produce soluble gp42 that mediates HLA class II immune evasion. J Virol. 2005, 79: 841-852. 10.1128/JVI.79.2.841-852.2005. Chackerian B: Virus-like particles: flexible platforms for vaccine development. Expert Rev Vaccines. 2007, 6: 381-390. 10.1586/14760584.6.3.381. Ramqvist T, Andreasson K, Dalianis T: Vaccination, immune and gene therapy based on virus-like particles against viral infections and cancer. Expert Opin Biol Ther. 2007, 7: 997-1007. 10.1517/14712598.7.7.997. Dargan DJ, Patel AH, Subak-Sharpe JH: PREPs: herpes simplex virus type 1-specific particles produced by infected cells when viral DNA replication is blocked. J Virol. 1995, 69: 4924-4932. Mocarski ES, Shenk T, Pass RF: Cytomegaloviruses. Fields Virology Volume 2. Edited by: Knipe DM, Howley PM, Griffin DE, Lamb RA, Martin MA, Roizman B, Straus SE. 2007, Philadelphia: Lippincott Williams &Wilkins, 2: 2701-2772. 5 Feederle R, Shannon-Lowe C, Baldwin G, Delecluse HJ: Defective infectious particles and rare packaged genomes produced by cells carrying terminal-repeat-negative Epstein-Barr virus. J Virol. 2005, 79: 7641-7647. 10.1128/JVI.79.12.7641-7647.2005. Adhikary D, Behrends U, Feederle R, Delecluse HJ, Mautner J: Standardized and highly efficient expansion of Epstein-Barr virus-specific CD4+ T cells by using virus-like particles. J Virol. 2008, 82: 3903-3911. 10.1128/JVI.02227-07. Tanaka M, Kagawa H, Yamanashi Y, Sata T, Kawaguchi Y: Construction of an excisable bacterial artificial chromosome containing a full-length infectious clone of herpes simplex virus type 1: viruses reconstituted from the clone exhibit wild-type properties in vitro and in vivo. J Virol. 2003, 77: 1382-1391. 10.1128/JVI.77.2.1382-1391.2003. Tischer BK, Kaufer BB, Sommer M, Wussow F, Arvin AM, Osterrieder N: A self-excisable infectious bacterial artificial chromosome clone of varicella-zoster virus allows analysis of the essential tegument protein encoded by ORF9. J Virol. 2007, 81: 13200-13208. 10.1128/JVI.01148-07. Zhou F, Li Q, Wong SW, Gao SJ: Autoexcision of bacterial artificial chromosome facilitated by terminal repeat-mediated homologous recombination: a novel approach for generating traceless genetic mutants of herpesviruses. J Virol. 2010, 84: 2871-2880. 10.1128/JVI.01734-09. Granato M, Feederle R, Farina A, Gonnella R, Santarelli R, Hub B, Faggioni A, Delecluse HJ: Deletion of Epstein-Barr virus BFLF2 leads to impaired viral DNA packaging and primary egress as well as to the production of defective viral particles. J Virol. 2008, 82: 4042-4051. 10.1128/JVI.02436-07. Farina A, Feederle R, Raffa S, Gonnella R, Santarelli R, Frati L, Angeloni A, Torrisi MR, Faggioni A, Delecluse HJ: BFRF1 of Epstein-Barr virus is essential for efficient primary viral envelopment and egress. J Virol. 2005, 79: 3703-3712. 10.1128/JVI.79.6.3703-3712.2005. Murata T, Isomura H, Yamashita Y, Toyama S, Sato Y, Nakayama S, Kudoh A, Iwahori S, Kanda T, Tsurumi T: Efficient production of infectious viruses requires enzymatic activity of Epstein-Barr virus protein kinase. Virology. 2009, 389: 75-81. 10.1016/j.virol.2009.04.007. Meng Q, Hagemeier SR, Kuny CV, Kalejta RF, Kenney SC: Simian virus 40 T/t antigens and lamin A/C small interfering RNA rescue the phenotype of an Epstein-Barr virus protein kinase (BGLF4) mutant. J Virol. 2010, 84: 4524-4533. 10.1128/JVI.02456-09. Feederle R, Bannert H, Lips H, Muller-Lantzsch N, Delecluse HJ: The Epstein-Barr virus alkaline exonuclease BGLF5 serves pleiotropic functions in virus replication. J Virol. 2009, 83: 4952-4962. 10.1128/JVI.00170-09. Neuhierl B, Delecluse HJ: The Epstein-Barr virus BMRF1 gene is essential for lytic virus replication. J Virol. 2006, 80: 5078-5081. 10.1128/JVI.80.10.5078-5081.2006. Nakayama S, Murata T, Murayama K, Yasui Y, Sato Y, Kudoh A, Iwahori S, Isomura H, Kanda T, Tsurumi T: Epstein-Barr virus polymerase processivity factor enhances BALF2 promoter transcription as a coactivator for the BZLF1 immediate-early protein. J Biol Chem. 2009, 284: 21557-21568. 10.1074/jbc.M109.015685. Feederle R, Neuhierl B, Baldwin G, Bannert H, Hub B, Mautner J, Behrends U, Delecluse HJ: Epstein-Barr virus BNRF1 protein allows efficient transfer from the endosomal compartment to the nucleus of primary B lymphocytes. J Virol. 2006, 80: 9435-9443. 10.1128/JVI.00473-06. Yu X, Wang Z, Mertz JE: ZEB1 regulates the latent-lytic switch in infection by Epstein-Barr virus. PLoS Pathog. 2007, 3: e194-10.1371/journal.ppat.0030194. Kelly GL, Milner AE, Tierney RJ, Croom-Carter DS, Altmann M, Hammerschmidt W, Bell AI, Rickinson AB: Epstein-Barr virus nuclear antigen 2 (EBNA2) gene deletion is consistently linked with EBNA3A, -3B, and -3C expression in Burkitt's lymphoma cells and with increased resistance to apoptosis. J Virol. 2005, 79: 10709-10717. 10.1128/JVI.79.16.10709-10717.2005. Skalska L, White RE, Franz M, Ruhmann M, Allday MJ: Epigenetic repression of p16(INK4A) by latent Epstein-Barr virus requires the interaction of EBNA3A and EBNA3C with CtBP. PLoS Pathog. 2010, 6: e1000951-10.1371/journal.ppat.1000951. Hertle ML, Popp C, Petermann S, Maier S, Kremmer E, Lang R, Mages J, Kempkes B: Differential gene expression patterns of EBV infected EBNA-3A positive and negative human B lymphocytes. PLoS Pathog. 2009, 5: e1000506-10.1371/journal.ppat.1000506. Maruo S, Wu Y, Ishikawa S, Kanda T, Iwakiri D, Takada K: Epstein-Barr virus nuclear protein EBNA3C is required for cell cycle progression and growth maintenance of lymphoblastoid cells. Proc Natl Acad Sci USA. 2006, 103: 19500-19505. 10.1073/pnas.0604919104. Dirmeier U, Neuhierl B, Kilger E, Reisbach G, Sandberg ML, Hammerschmidt W: Latent membrane protein 1 is critical for efficient growth transformation of human B cells by epstein-barr virus. Cancer Res. 2003, 63: 2982-2989. Mancao C, Hammerschmidt W: Epstein-Barr virus latent membrane protein 2A is a B-cell receptor mimic and essential for B-cell survival. Blood. 2007, 110: 3715-3721. 10.1182/blood-2007-05-090142. Feederle R, Delecluse HJ: Low level of lytic replication in a recombinant Epstein-Barr virus carrying an origin of replication devoid of BZLF1-binding sites. J Virol. 2004, 78: 12082-12084. 10.1128/JVI.78.21.12082-12084.2004. Chau CM, Zhang XY, McMahon SB, Lieberman PM: Regulation of Epstein-Barr virus latency type by the chromatin boundary factor CTCF. J Virol. 2006, 80: 5723-5732. 10.1128/JVI.00025-06. Hutzinger R, Feederle R, Mrazek J, Schiefermeier N, Balwierz PJ, Zavolan M, Polacek N, Delecluse HJ, Huttenhofer A: Expression and processing of a small nucleolar RNA from the Epstein-Barr virus genome. PLoS Pathog. 2009, 5: e1000547-10.1371/journal.ppat.1000547. Delecluse HJ, Pich D, Hilsendegen T, Baum C, Hammerschmidt W: A first-generation packaging cell line for Epstein-Barr virus-derived vectors. Proc Natl Acad Sci USA. 1999, 96: 5188-5193. 10.1073/pnas.96.9.5188.