Abstract:
:Engineered alphavirus vectors expressing reporters of infection have been used for a number of years due to their relatively low costs for analysis of virus replication and the capacity to utilize imaging systems for longitudinal measurements of growth within single animals. In general, these vectors have been derived from Old World alphaviruses using a second viral subgenomic promoter to express the transgenes, placed either immediately after the nonstructural proteins or at the 3' end of the viral coding sequences. However, the relevance of these vectors to natural infections is questionable, as they have not been rigorously tested for virulence in vivo in comparison with parental viruses or for the retention of the reporter during replication. Here, we report construction of new expression vectors for two Old World arthritogenic alphaviruses (Sindbis and Chikungunya viruses) and two New World encephalitic alphaviruses (eastern and Venezuelan equine encephalitis viruses) based upon either fusion of the reporter protein in frame within nonstructural protein 3 (nsP3) or insertion of the reporter as a cleavable element between the capsid and PE2 structural proteins. We have compared these with a traditional 3' double subgenomic promoter virus expressing either a large, firefly luciferase (fLuc; 1,650 nucleotides), or small, NanoLuc (nLuc; 513 nucleotides), luminescent reporter protein. Results indicate that the nLuc is substantially more stable than fLuc during repeated rounds of infection regardless of the transgene location. However, the capsid-PE2 insertion and nsP3 fusion viruses exhibit the most authentic mimicking of parental virus infection regardless of expressed protein. IMPORTANCE As more antiviral therapeutics and vaccines are developed, rapid and accurate in vivo modeling of their efficacy will be required. However, current alphavirus vectors expressing reporters of infection have not been extensively tested for accurate mimicking of the infection characteristics of unmodified parental viruses. Additionally, use of in vivo imaging systems detecting light emitted from luciferase reporters can significantly decrease costs associated with efficacy studies by minimizing numbers of animals. Herein we report development and testing of new expression vectors for Sindbis, Chikungunya, and eastern and Venezuelan equine encephalitis viruses and demonstrate that a small (∼500-nucleotide) reporter gene (NanoLuc; Promega) is very stable and causes a disease severity similar to that caused by unmodified parental viruses. In contrast, expression of larger reporters is very rapidly lost with virus replication and can be significantly attenuating. The utility of NanoLuc for in vivo imaging is also demonstrated.
journal_name
J Viroljournal_title
Journal of virologyauthors
Sun C,Gardner CL,Watson AM,Ryman KD,Klimstra WBdoi
10.1128/JVI.02990-13subject
Has Abstractpub_date
2014-02-01 00:00:00pages
2035-46issue
4eissn
0022-538Xissn
1098-5514pii
JVI.02990-13journal_volume
88pub_type
杂志文章abstract::DNA replication intermediates of three plasmids containing all or part of a modified Epstein-Barr virus cis-acting plasmid maintenance region (oriP) were examined to further investigate oriP function. Replication intermediates were analyzed in vivo and in vitro by neutral-neutral two-dimensional gel electrophoresis. T...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.67.3.1739-1745.1993
更新日期:1993-03-01 00:00:00
abstract::Intragastric inoculation with whole-virion vaccine of inactivated influenza virus resulted in production of hemagglutinin (HA)-specific immunoglobulin A (IgA) and IgG both in lung lavage fluids and in serum samples of mice. HA-specific IgA was the predominant isotypic antibody secreted in the lung lavage fluids (avera...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.61.7.2150-2154.1987
更新日期:1987-07-01 00:00:00
abstract::Apoptin, a viral death protein derived from chicken anemia virus, displays a number of tumor-specific behaviors. In particular, apoptin is phosphorylated, translocates to the nucleus, and induces apoptosis specifically in tumor or transformed cells, whereas it is nonphosphorylated and remains primarily inactive in the...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.78.18.9965-9976.2004
更新日期:2004-09-01 00:00:00
abstract::The effectiveness of attenuated poliovirus vaccines when given orally to induce both systemic and mucosal immune responses against poliovirus has resulted in an effort to develop poliovirus-based vectors to express foreign proteins. We have previously described the construction of poliovirus genomes (referred to as re...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.70.4.2643-2649.1996
更新日期:1996-04-01 00:00:00
abstract::We have developed a high-throughput sequencing (HTS) workflow for investigating paramyxovirus transcription and replication. We show that sequencing of oligo(dT)-selected polyadenylated mRNAs, without considering the orientation of the RNAs from which they had been generated, cannot accurately be used to analyze the a...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.00571-19
更新日期:2019-08-13 00:00:00
abstract::HLA-B*57 is strongly associated with immune control of HIV and delayed AIDS progression. The closely related, but less protective, HLA-B*58:01 presents similar epitopes, but HLA-B*58:01(+) individuals do not generate CD8(+) T cells targeting the KF11-Gag epitope, which has been linked to low viremia. Here we show that...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.01122-13
更新日期:2013-10-01 00:00:00
abstract::Measles virus (MV) infection causes an acute childhood disease, associated in certain cases with infection of the central nervous system and development of a severe neurological disease. We have generated transgenic mice ubiquitously expressing the human protein SLAM (signaling lymphocytic activation molecule), or CD1...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.00209-06
更新日期:2006-07-01 00:00:00
abstract::The copy choice model for the generation of defective interfering (DI) particles of vesicular stomatitis virus suggests that during replication the polymerase prematurely terminates, moves with the nascent daughter strand to another site on the same or a different template molecule, and resumes elongation of the nasce...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.51.2.515-521.1984
更新日期:1984-08-01 00:00:00
abstract::Monoclonal antibodies produced against the prototype cell-adapted Wyoming strain of equine infectious anemia virus (EIAV), a lentivirus, were studied for reactivity with the homologous prototype and 16 heterologous isolates. Eighteen hybridomas producing monoclonal antibodies (MAbs) were isolated. Western blot (immuno...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.61.10.2956-2961.1987
更新日期:1987-10-01 00:00:00
abstract::We have expressed the human immunodeficiency virus type 1 (HIV-1) protease (PR) in bacteria as a Gag-PR polyprotein (J. Luban and S.P. Goff, J. Virol. 65:3203-3212, 1991). The protein displays enzymatic activity, cleaving the Gag polyprotein precursor Pr55gag to the expected products. The PR enzyme is only active as a...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.67.6.3630-3634.1993
更新日期:1993-06-01 00:00:00
abstract::The NS5A protein of hepatitis C virus (HCV) plays an important but undefined role in viral RNA replication. NS5A has been proposed to be a three-domain protein, and the crystal structure of the well-conserved amino-terminal domain I has been determined. The remaining two domains of NS5A, designated domains II and III,...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.00328-07
更新日期:2008-02-01 00:00:00
abstract::The great majority of viral mRNAs in mouse C127 cells transformed by bovine papillomavirus type 1 (BPV) have a common 3' end at the early polyadenylation site which is 23 nucleotides (nt) downstream of a canonical poly(A) consensus signal. Twenty percent of BPV mRNA from productively infected cells bypasses the early ...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.67.12.7705-7710.1993
更新日期:1993-12-01 00:00:00
abstract::We used 18 monoclonal antibodies against B19 parvovirus to identify neutralizing epitopes on the viral capsid. Of the 18 antibodies, 9 had in vitro neutralizing activity in a bone marrow colony culture assay. The overlapping polypeptide fragments spanning the B19 structural proteins were produced in a pMAL-c Escherich...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.65.12.7056-7060.1991
更新日期:1991-12-01 00:00:00
abstract::The transmission bottleneck is defined as the number of viral particles that transmit from one host to establish an infection in another. Genome sequence data have been used to evaluate the size of the transmission bottleneck between humans infected with the influenza virus; however, the methods used to make these est...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.00014-20
更新日期:2020-06-16 00:00:00
abstract::The process of Epstein-Barr virus (EBV)-induced transformation of human B lymphocytes results in a cell line that is a mixture of latently and lytically infected cells, with the lytic cells composing roughly 5% to less than 0.0001% of the overall population. A set of nine normal lymphoblastoid cell lines that span a 1...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.64.1.437-444.1990
更新日期:1990-01-01 00:00:00
abstract::The UL3 protein of equine herpesvirus 1 (EHV-1) KyA strain is a homolog of the ICP27 alpha regulatory protein of herpes simplex virus type 1 (HSV-1) and the ORF 4 protein of varicella-zoster virus. To characterize the regulatory function of the UL3 gene product, a UL3 gene expression vector (pSVUL3) and a vector expre...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.69.5.2786-2793.1995
更新日期:1995-05-01 00:00:00
abstract::Since dendritic cells may play a key role in defense against influenza virus infection, we examined the effects of recombinant hemagglutinin (HA) proteins derived from mouse-adapted H1N1 (A/WSN/1933), swine-origin 2009 pandemic H1N1 (A/Texas/05/2009), and highly pathogenic avian influenza H5N1 (A/Thailand/KAN-1/2004) ...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.01316-10
更新日期:2010-11-01 00:00:00
abstract::Defined segments of the cloned foot-and-mouth disease virus genome corresponding to all parts of the coding region were expressed in Escherichia coli as fusions to the N-terminal part of the MS2-polymerase gene under the control of the inducible lambda PL promoter. All constructs yielded large amounts of proteins, whi...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.57.3.983-991.1986
更新日期:1986-03-01 00:00:00
abstract::Widespread distribution of highly pathogenic avian H5N1 influenza viruses in domesticated and wild birds continues to pose a threat to public health, as interspecies transmission of virus has resulted in increasing numbers of human disease cases. Although the pathogenic mechanism(s) of H5N1 influenza viruses has not b...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.02144-08
更新日期:2009-06-01 00:00:00
abstract::The helicase-like 1a and polymerase-like 2a proteins of brome mosaic virus (BMV) are required for viral RNA replication in vivo, are present in membrane-bound viral RNA polymerase extracts, and share conservation with the many other members of the alphavirus-like superfamily. To better understand BMV RNA replication a...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.70.12.8908-8916.1996
更新日期:1996-12-01 00:00:00
abstract::The papillomavirus E2 protein is a critical viral regulatory protein with transcription, DNA replication, and genome maintenance functions. We have previously identified the cellular bromodomain protein Brd4 as a major E2-interacting protein and established that it participates in tethering bovine papillomavirus type ...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.00447-07
更新日期:2007-09-01 00:00:00
abstract::Worldwide amphibian population declines have been ascribed to global warming, increasing pollution levels, and other factors directly related to human activities. These factors may additionally be favoring the emergence of novel pathogens. In this report, we have determined the complete genome sequence of the emerging...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.07108-11
更新日期:2012-04-01 00:00:00
abstract::Sensory neurons within trigeminal ganglia (TG) are the primary site for bovine herpesvirus 1 (BHV-1) latency. During latency, viral gene expression is restricted to the latency-related (LR) gene and the open reading frame ORF-E. We previously constructed an LR mutant virus that expresses LR RNA but not any of the know...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.01009-09
更新日期:2009-09-01 00:00:00
abstract::The yield of G418-resistant Huh7 cell clones bearing subgenomic dicistronic GB virus B (GBV-B) is significantly affected by the insertion of a portion of the viral core gene between the GBV-B 5' untranslated region and the exogenous neomycin phosphotransferase selector gene (A. De Tomassi, M. Pizzuti, R. Graziani, A. ...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/jvi.77.13.7502-7509.2003
更新日期:2003-07-01 00:00:00
abstract::A lysate from hypotonically swollen polyoma-infected BALB/3T3 cells incorporated labeled deoxynucleotide triphosphates into both viral and cellular DNAs. The incorporation was stimulated by the presence of ATP, deoxynucleotide triphosphates, thiols, and magnesium ions. Strong inhibition of incorporation was observed w...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.13.1.125-139.1974
更新日期:1974-01-01 00:00:00
abstract::To date, no vaccine that is safe and effective against herpes simplex virus 2 (HSV-2) disease has been licensed. In this study, we evaluated a DNA prime-formalin-inactivated-HSV-2 (FI-HSV2) boost vaccine approach in the guinea pig model of acute and recurrent HSV-2 genital disease. Five groups of guinea pigs were immu...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.02521-10
更新日期:2011-04-01 00:00:00
abstract::Tick-borne encephalitis virus (TBEV), a major tick-borne viral pathogen of humans, is known to cause neurological diseases such as meningitis, encephalitis, and meningoencephalitis. However, the life cycle and pathogenesis of TBEV are not well understood. Here, we show that the knockdown or knockout of ADAM15 (a disin...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.01926-20
更新日期:2021-01-28 00:00:00
abstract:UNLABELLED:Human T-cell lymphotropic virus type 1 (HTLV-1) and HTLV-2 encode auxiliary proteins that play important roles in viral replication, viral latency, and immune escape. The presence of auxiliary protein-encoding open reading frames (ORFs) in HTLV-3, the latest HTLV to be discovered, is unknown. Simian T-cell l...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.02150-14
更新日期:2015-01-15 00:00:00
abstract::The yeast two-hybrid system was used to catalog all detectable interactions among the P2 nonstructural cleavage products of poliovirus type 1 (Mahoney). Evidence has been obtained for specific associations among 2A(pro), 2BC, 2C, and 2B. Specifically, 2A(pro) can interact with itself and 2BC and its cleavage products ...
journal_title:Journal of virology
pub_type: 杂志文章
doi:10.1128/JVI.72.2.1297-1307.1998
更新日期:1998-02-01 00:00:00
abstract::Plasma human immunodeficiency virus type 1 (HIV-1) turnover and kinetics were studied in children aged 15 days to 2 years following the initiation of a triple antiretroviral drug regimen consisting of zidovudine, lamivudine, and nevirapine. HIV-1 turnover was at least as rapid as that previously described in adults; t...
journal_title:Journal of virology
pub_type: 临床试验,杂志文章
doi:10.1128/JVI.73.1.362-367.1999
更新日期:1999-01-01 00:00:00