Abstract
Using the RNA replication machinery of Japanese encephalitis virus (JEV), the authors have established and characterized three strategies for the expression of foreign genes. Initially, ≈11 kb genomic RNA was engineered to express heterologous genes of various sizes by preferentially inserting a new cistron at the beginning of the 3′ nontranslated variable region. RNA transfection yielded recombinant viruses that initiated foreign gene expression after infecting permissive cells. JEV was capable of packaging recombinant genomes as large as ≈15 kb. However, larger genome size was inversely correlated with RNA replication efficiency and cytopathogenicity, with no significant change in infectivity. Second, a variety of self-replicating propagation-deficient viral replicons were constructed by introducing one to three in-frame deletions into the ectodomains of all the structural proteins of JEV. These replicons displayed a spectrum of RNA replication efficiency upon transfection, suggesting that remnant transmembrane domains play a suppressive role in this process. Third, the authors generated a panel of stable packaging cell lines (PCLs) providing all three JEV structural proteins in trans. These PCLs efficiently packaged viral replicon RNAs into single-round infectious viral replicon particles. These JEV-based virus/vector systems may provide useful tools for a variety of biological applications, including foreign gene expression, antiviral compound screening, and genetic immunization.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Agapov EV, Frolov I, Linderbach BD, Pragai BM, Schlesinger S, Rice CM (1998). Noncytopathic Sindbis virus RNA replicons for heterologous gene expression. Proc Natl Acad Sci U S A 95: 12989–12994.
Alvarez DE, Lodeiro MF, Luduena SJ, Pietrasanta LI, Gamarnik AV (2005). Long-range RNA-RNA interactions circularize the dengue virus genome. J Virol 79: 6631–6643.
Bonaldo MC, Garratt RC, Caufour PS, Freire MS, Rodrigues MM, Nussenzweig RS, Galler R (2002). Surface expression of an immunodominant malaria protein B cell epitope by yellow fever virus. J Mol Biol 315: 873–885.
Bonaldo MC, Garratt RC, Marchevsky RS, Coutinho ES, Jabor AV, Almeida LF, Yamamura AM, Duarte AS, Oliveira PJ, Lizeu JO, Camacho LA, Freire MS, Galler R (2005). Attenuation of recombinant yellow fever 17D viruses expressing foreign protein epitopes at the surface. J Virol 79: 8602–8613.
Bredenbeek PJ, Kooi EA, Lindenbach B, Huijkman N, Rice CM, Spaan WJ (2003). A stable full-length yellow fever virus cDNA clone and the role of conserved RNA elements in flavivirus replication. J Gen Virol 84: 1261–1268.
Bredenbeek PJ, Molenkamp R, Spaan WJ, Deubel V, Marianneau P, Salvato MS, Moshkoff D, Zapata J, Tikhonov I, Patterson J, Carrion R, Ticer A, Brasky K, Lukashevich IS (2006). A recombinant yellow fever 17D vaccine expressing Lassa virus glycoproteins. Virology 345: 299–304.
Corver J, Lenches E, Smith K, Robison RA, Sando T, Strauss EG, Strauss JH (2003). Fine mapping of a cis-acting sequence element in yellow fever virus RNA that is required for RNA replication and cyclization. J Virol 77: 2265–2270.
Fayzulin R, Scholle F, Petrakova O, Frolov I, Mason PW (2006). Evaluation of replicative capacity and genetic stability of West Nile virus replicons using highly efficient packaging cell lines. Virology 351: 196–209.
Flotte TR, Afione SA, Solow R, Drumm ML, Markakis D, Guggino WB, Zeitlin PL, Carter BJ (1993). Expression of the cystic fibrosis transmembrane conductance regulator from a novel adeno-associated virus promoter. J Biol Chem 268: 3781–3790.
Gehrke R, Ecker M, Aberle SW, Allison SL, Heinz FX, Mandl CW (2003). Incorporation of tick-borne encephalitis virus replicons into virus-like particles by a packaging cell line. J Virol 77: 8924–8933.
Gehrke R, Heinz FX, Davis NL, Mandl CW (2005). Heterologous gene expression by infectious and replicon vectors derived from tick-borne encephalitis virus and direct comparison of this flavivirus system with an alphavirus replicon. J Gen Virol 86: 1045–1053.
Hahn CS, Hahn YS, Rice CM, Lee E, Dalgarno L, Strauss EG, Strauss JH (1987). Conserved elements in the 3′ untranslated region of flavivirus RNAs and potential cyclization sequences. J Mol Biol 198: 33–41.
Harvey TJ, Liu WJ, Wang XJ, Linedale R, Jacobs M, Davidson A, Le TT, Anraku I, Suhrbier A, Shi PY, Khromykh AA (2004). Tetracycline-inducible packaging cell line for production of flavivirus replicon particles. J Virol 78: 531–538.
Hennessy S, Liu Z, Tsai TF, Strom BL, Wan CM, Liu HL, Wu TX, Yu HJ, Liu QM, Karabatsos N, Bilker WB, Halstead SB (1996). Effectiveness of live-attenuated Japanese encephalitis vaccine (SA14-14-2): a case control study. Lancet 347: 1583–1586.
Jones CT, Patkar CG, Kuhn RJ (2005). Construction and applications of yellow fever virus replicons. Virology 331: 247–259.
Kaufman RJ (1999). Stress signaling from the lumen of the endoplasmic reticulum: coordination of gene transcriptional and translational controls. Genes Dev 13: 1211–1233.
Khromykh AA, Meka H, Guyatt KJ, Westaway EG (2001). Essential role of cyclization sequences in flavivirus RNA replication. J Virol 75: 6719–6728.
Khromykh AA, Varnavski AN, Westaway EG (1998). Encapsidation of the flavivirus Kunjin replicon RNA by using a complementation system providing Kunjin virus structural proteins in trans. J Virol 72: 5967–5977.
Khromykh AA, Westaway EG (1997). Subgenomic replicons of the flavivirus Kunjin: construction and applications. J Virol 71: 1497–1505.
Lai CJ, Monath TP (2003). Chimeric flaviviruses: novel vaccines against dengue fever, tick-borne encephalitis, and Japanese encephalitis. Adv Virus Res 61: 469–509.
Lindenbach BD, Rice CM (2001). Flaviviridae: The viruses and their replication. In: Fields virology, 4th ed. Knipe DM, Howley PM, Griffin DE, Lamb RA, Martin MA, Roizman B, Straus SE (eds). Philadelphia: Lippincott Williams & Wilkins Publishers, pp 991–1041.
Lo MK, Tilgner M, Bernard KA, Shi PY (2003). Functional analysis of mosquito-borne flavivirus conserved sequence elements within 3′ untranslated region of West Nile virus by use of a reporting replicon that differentiates between viral translation and RNA replication. J Virol 77: 10004–10014.
Markoff L (2003). 5′- and 3′-noncoding regions in flavivirus RNA. Adv Virus Res 59: 177–228.
Mason PW, Shustov AV, Frolov I (2006). Production and characterization of vaccines based on flaviviruses defective in replication. Virology 351: 432–443.
Molenkamp R, Kooi EA, Lucassen MA, Greve S, Thijssen JC, Spaan WJ, Bredenbeek PJ (2003). Yellow fever virus replicons as an expression system for hepatitis C virus structural proteins. J Virol 77: 1644–1648.
Monath TP (2003). Yellow fever vaccine, 4th ed. Philadelphia: WB Saunders.
Pahl HL (1999). Signal transduction from the endoplasmic reticulum to the cell nucleus. Physiol Rev 79: 683–701.
Pang X, Zhang M, Dayton AI (2001). Development of dengue virus replicons expressing HIV-1 gp120 and other heterologous genes: a potential future tool for dual vaccination against dengue virus and HIV. BMC Microbiol 1: 28–36.
Pierson TC, Diamond MS, Ahmed AA, Valentine LE, Davis CW, Samuel MA, Hanna SL, Puffer BA, Doms RW (2005). An infectious West Nile virus that expresses a GFP reporter gene. Virology 334: 28–40.
Scholle F, Girard YA, Zhao Q, Higgs S, Mason PW (2004). trans-Packaged West Nile virus-like particles: infectious properties in vitro and in infected mosquito vectors. J Virol 78: 11605–11614.
Shi PY, Tilgner M, Lo MK (2002). Construction and characterization of subgenomic replicons of New York strain of West Nile virus. Virology 296: 219–233.
Solomon T (2003). Recent advances in Japanese encephalitis. J NeuroVirol 9: 274–283.
Su HL, Liao CL, Lin YL (2002). Japanese encephalitis virus infection initiates endoplasmic reticulum stress and an unfolded protein response. J Virol 76: 4162–4171.
Varnavski AN, Khromykh AA (1999). Noncytopathic flavivirus replicon RNA-based system for expression and delivery of heterologous genes. Virology 255: 366–375.
Wang JJ, Liao CL, Chiou YW, Chiou CT, Huang YL, Chen LK (1997). Ultrastructure and localization of E proteins in cultured neuron cells infected with Japanese encephalitis virus. Virology 238: 30–39.
Winer J, Jung CK, Shackel I, Williams PM (1999). Development and validation of real-time quantitative reverse transcriptase-polymerase chain reaction for monitoring gene expression in cardiac myocytes in vitro. Anal Biochem 270: 41–49.
Xin YY, Ming ZG, Peng GY, Jian A, Min LH (1988). Safety of a live-attenuated Japanese encephalitis virus vaccine (SA14-14-2) for children. Am J Trop Med Hyg 39: 214–217.
You S, Falgout B, Markoff L, Padmanabhan R (2001). In vitro RNA synthesis from exogenous dengue viral RNA templates requires long range interactions between 5′- and 3′-terminal regions that influence RNA structure. J Biol Chem 276: 15581–15591.
Yun SI, Kim SY, Rice CM, Lee YM (2003). Development and application of areverse genetics system for Japanese encephalitis virus. J Virol 77: 6450–6465.
Author information
Authors and Affiliations
Corresponding author
Additional information
Supplemental materials for this article are provided separately.
This work was supported by a Korea Research Foundation grant (KRF-2001-042-D00071), Republic of Korea.
Rights and permissions
About this article
Cite this article
Yun, SI., Choi, YJ., Yu, XF. et al. Engineering the Japanese encephalitis virus RNA genome for the expression of foreign genes of various sizes: Implications for packaging capacity and RNA replication efficiency. Journal of NeuroVirology 13, 522–535 (2007). https://doi.org/10.1080/13550280701684651
Received:
Revised:
Accepted:
Issue Date:
DOI: https://doi.org/10.1080/13550280701684651