Scott J Goebel

New York State Department of Health, New York City, NY, United States

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Publications (20)78.65 Total impact

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    ABSTRACT: The coronavirus nucleocapsid (N) protein plays an essential role in virion assembly via interactions with the large, positive-strand RNA viral genome and the carboxy-terminal endodomain of the membrane protein (M). To learn about the functions of N protein domains in the coronavirus mouse hepatitis virus (MHV), we replaced the MHV N gene with its counterpart from the closely related bovine coronavirus (BCoV). The resulting viral mutant was severely defective, even though individual domains of the N protein responsible for N-RNA, N-M, or N-N interactions were completely interchangeable between BCoV and MHV. The lesion in the BCoV N substitution mutant could be compensated for by reverting mutations in the central, serine- and arginine-rich (SR) domain of the N protein. Surprisingly, a second class of reverting mutations were mapped to the amino terminus of a replicase subunit, nonstructural protein 3 (nsp3). A similarly defective MHV N mutant bearing an insertion of the SR region from the severe acute respiratory syndrome coronavirus N protein was rescued by the same two classes of reverting mutations. Our genetic results were corroborated by the demonstration that the expressed amino-terminal segment of nsp3 bound selectively to N protein from infected cells, and this interaction was RNA independent. Moreover, we found a direct correlation between the N-nsp3 interaction and the ability of N protein to stimulate the infectivity of transfected MHV genomic RNA (gRNA). Our results suggest a role for this previously unknown N-nsp3 interaction in the localization of genomic RNA to the replicase complex at an early stage of infection.
    Journal of Virology 10/2010; 84(19):10276-88. · 5.08 Impact Factor
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    ABSTRACT: The type I interferon (IFN) response plays an essential role in the control of in vivo infection by the coronavirus mouse hepatitis virus (MHV). However, in vitro, most strains of MHV are largely resistant to the action of this cytokine, suggesting that MHV encodes one or more functions that antagonize or evade the IFN system. A particular strain of MHV, MHV-S, exhibited orders-of-magnitude higher sensitivity to IFN than prototype strain MHV-A59. Through construction of interstrain chimeric recombinants, the basis for the enhanced IFN sensitivity of MHV-S was found to map entirely to the region downstream of the spike gene, at the 3' end of the genome. Sequence analysis revealed that the major difference between the two strains in this region is the absence of gene 5a from MHV-S. Creation of a gene 5a knockout mutant of MHV-A59 demonstrated that a major component of IFN resistance maps to gene 5a. Conversely, insertion of gene 5a, or its homologs from related group 2 coronaviruses, at an upstream genomic position in an MHV-A59/S chimera restored IFN resistance. This is the first demonstration of a coronavirus gene product that can protect that same virus from the antiviral state induced by IFN. Neither protein kinase R, which phosphorylates eukaryotic initiation factor 2, nor oligoadenylate synthetase, which activates RNase L, was differentially activated in IFN-treated cells infected with MHV-A59 or MHV-S. Thus, the major IFN-induced antiviral activities that are specifically inhibited by MHV, and possibly by other coronaviruses, remain to be identified.
    Journal of Virology 08/2010; 84(16):8262-74. · 5.08 Impact Factor
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    ABSTRACT: The upstream end of the 3' untranslated region (UTR) of the mouse hepatitis virus genome contains two essential and overlapping RNA secondary structures, a bulged stem-loop and a pseudoknot, which have been proposed to be elements of a molecular switch that is critical for viral RNA synthesis. It has previously been shown that a particular six-base insertion in loop 1 of the pseudoknot is extremely deleterious to the virus. We have now isolated multiple independent second-site revertants of the loop 1 insertion mutant, and we used reverse-genetics methods to confirm the identities of suppressor mutations that could compensate for the original insertion. The suppressors were localized to two separate regions of the genome. Members of one class of suppressor were mapped to the portions of gene 1 that encode nsp8 and nsp9, thereby providing the first evidence for specific interactions between coronavirus replicase gene products and a cis-acting genomic RNA element. The second class of suppressor was mapped to the extreme 3' end of the genome, a result which pointed to the existence of a direct base-pairing interaction between loop 1 of the pseudoknot and the genomic terminus. The latter finding was strongly supported by phylogenetic evidence and by the construction of a deletion mutant that reduced the 3' UTR to its minimal essential elements. Taken together, the interactions revealed by the two classes of suppressors suggest a model for the initiation of coronavirus negative-strand RNA synthesis.
    Journal of Virology 03/2008; 82(3):1214-28. · 5.08 Impact Factor
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    ABSTRACT: An unknown virus was isolated from a lung biopsy sample and multiple other samples from a patient who developed a lethal case of pneumonia following a peripheral blood stem cell transplant. A random PCR-based molecular screening method was used to identify the infectious agent as avian paramyxovirus 1 (APMV-1; a group encompassing Newcastle disease virus), which is a highly contagious poultry pathogen that has only rarely been found in human infections. Immunohistochemical analysis confirmed the presence of APMV-1 antigen in sloughed alveolar cells in lung tissue from autopsy. Sequence from the human isolate showed that it was most closely related to virulent pigeon strains of APMV-1. This is the most completely documented case of a systemic human infection caused by APMV-1 and is the first report of an association between this virus and a fatal disease in a human.
    Journal of Virology 12/2007; 81(22):12709-14. · 5.08 Impact Factor
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    ABSTRACT: The 3' cis-acting element for mouse hepatitis virus (MHV) RNA synthesis resides entirely within the 301-nucleotide 3' untranslated region (3' UTR) of the viral genome and consists of three regions. Encompassing the upstream end of the 3' UTR are a bulged stem-loop and an overlapping RNA pseudoknot, both of which are essential to MHV and common to all group 2 coronaviruses. At the downstream end of the genome is the minimal signal for initiation of negative-strand RNA synthesis. Between these two ends is a hypervariable region (HVR) that is only poorly conserved between MHV and other group 2 coronaviruses. Paradoxically, buried within the HVR is an octanucleotide motif (oct), 5'-GGAAGAGC-3', which is almost universally conserved in coronaviruses and is therefore assumed to have a critical biological function. We conducted an extensive mutational analysis of the HVR. Surprisingly, this region tolerated numerous deletions, rearrangements, and point mutations. Most striking, a mutant deleted of the entire HVR was only minimally impaired in tissue culture relative to the wild type. By contrast, the HVR deletion mutant was highly attenuated in mice, causing no signs of clinical disease and minimal weight loss compared to wild-type virus. Correspondingly, replication of the HVR deletion mutant in the brains of mice was greatly reduced compared to that of the wild type. Our results show that neither the HVR nor oct is essential for the basic mechanism of MHV RNA synthesis in tissue culture. However, the HVR appears to play a significant role in viral pathogenesis.
    Journal of Virology 03/2007; 81(3):1274-87. · 5.08 Impact Factor
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    ABSTRACT: Triaryl pyrazoline {[5-(4-chloro-phenyl)-3-thiophen-2-yl-4,5-dihydro-pyrazol-1-yl]-phenyl-methanone} inhibits flavivirus infection in cell culture. The inhibitor was identified through high-throughput screening of a compound library using a luciferase-expressing West Nile (WN) virus infection assay. The compound inhibited an epidemic strain of WN virus without detectable cytotoxicity (a 50% effective concentration of 28 microM and a compound concentration of >or=300 microM required to reduce 50% cell viability). Besides WN virus, the compound also inhibited other flaviviruses (dengue, yellow fever, and St. Louis encephalitis viruses), an alphavirus (Western equine encephalitis virus), a coronavirus (mouse hepatitis virus), and a rhabdovirus (vesicular stomatitis virus). However, the compound did not suppress an orthomyxovirus (influenza virus) or a retrovirus (human immunodeficiency virus type 1). Mode-of-action analyses in WN virus showed that the compound did not inhibit viral entry or virion assembly but specifically suppressed viral RNA synthesis. To examine the mechanism of inhibition of dengue virus, we developed two replicon systems for dengue type 1 virus: (i) a stable cell line that harbored replicons containing a luciferase reporter and a neomycin phosphotransferase selection marker and (ii) a luciferase-expressing replicon that could differentiate between viral translation and RNA replication. Analyses of the compound in the dengue type 1 virus replicon systems showed that it weakly suppressed viral translation but significantly inhibited viral RNA synthesis. Overall, the results demonstrate that triaryl pyrazoline exerts a broad spectrum of antiflavivirus activity through potent inhibition of viral RNA replication. This novel inhibitor could be developed for potential treatment of flavivirus infection.
    Antimicrobial Agents and Chemotherapy 04/2006; 50(4):1320-9. · 4.57 Impact Factor
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    Scott J Goebel, Jill Taylor, Paul S Masters
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    ABSTRACT: The 3' untranslated region (3' UTR) of the genome of the severe acute respiratory syndrome coronavirus can functionally replace its counterpart in the prototype group 2 coronavirus mouse hepatitis virus (MHV). By contrast, the 3' UTRs of representative group 1 or group 3 coronaviruses cannot operate as substitutes for the MHV 3' UTR.
    Journal of Virology 08/2004; 78(14):7846-51. · 5.08 Impact Factor
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    ABSTRACT: RNA virus genomes contain cis-acting sequence and structural elements that participate in viral replication. We previously identified a bulged stem-loop secondary structure at the upstream end of the 3' untranslated region (3' UTR) of the genome of the coronavirus mouse hepatitis virus (MHV). This element, beginning immediately downstream of the nucleocapsid gene stop codon, was shown to be essential for virus replication. Other investigators discovered an adjacent downstream pseudoknot in the 3' UTR of the closely related bovine coronavirus (BCoV). This pseudoknot was also shown to be essential for replication, and it has a conserved counterpart in every group 1 and group 2 coronavirus. In MHV and BCoV, the bulged stem-loop and pseudoknot are, in part, mutually exclusive, because of the overlap of the last segment of the stem-loop and stem 1 of the pseudoknot. This led us to hypothesize that they form a molecular switch, possibly regulating a transition occurring during viral RNA synthesis. We have now performed an extensive genetic analysis of the two components of this proposed switch. Our results define essential and nonessential components of these structures and establish the limits to which essential parts of each element can be destabilized prior to loss of function. Most notably, we have confirmed the interrelationship of the two putative switch elements. Additionally, we have identified a pseudoknot loop insertion mutation that appears to point to a genetic interaction between the pseudoknot and a distant region of the genome.
    Journal of Virology 02/2004; 78(2):669-82. · 5.08 Impact Factor
  • G P Johnson, S J Goebel, E Paoletti
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    ABSTRACT: This communication is intended as a single source update of the initial report (Goebel et al., 1990a,b) which described the complete DNA sequence of the vaccinia virus genome. We have integrated published information as well as unpublished data. Our understanding of the complexities of the genetic functional organization of poxviruses is increasing at a remarkable rate. While some previously unknown identities have since been elucidated, the fact that the majority of vaccinia-encoded gene products still lack assigned functions lends excitement to the immediate future of poxvirus research.
    Virology 11/1993; 196(2):381-401. · 3.37 Impact Factor
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    ABSTRACT: Recently, we have reported the complete nucleotide sequence of vaccinia virus (Goebel, S. J., Johnson, G. P., Perkus, M. E., Davis, S. W., Winslow, J. P., and Paoletti, E. 1990, Virology 179, 247-266). Approximately 2.2 kbp leftward of the large subunit of ribonucleotide reductase resides a 108-amino acid open reading frame, O2L (nt 62,851-62,528) with significant similarity to known glutaredoxins. The deduced amino acid sequence of open reading frame O2L is 28.7% identical to the yeast and Escherichia coli proteins and greater than 40% identical to various mammalian glutaredoxins. Similar patterns of hydrophobicity as well as alpha-helix and beta-sheet potentials suggest that O2L and the glutaredoxins share a similar secondary structure. Furthermore, a common function is inferred by the presence of a highly conserved redox-active site.
    Virology 04/1991; 181(1):378-81. · 3.37 Impact Factor
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    ABSTRACT: Each copy of the inverted terminal repeat of vaccinia virus consists of 8 kb of DNA containing 9 ORFS flanked near the terminus of the genome by 4 kb of repetitive DNA which in turn contains blocks of tandem repeats. Using plasmids containing repetitive DNA as the external arm, we have generated deletions at both the left and the right termini of the vaccinia genome. We report here the engineered deletion within a single vaccinia virus of 32.7 kb of DNA (including 38 ORFS) from the left terminus and 14.9 kb of DNA (including 17 ORFS) from the right terminus.
    Virology 02/1991; 180(1):406-10. · 3.37 Impact Factor
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    ABSTRACT: A gene encoding an 18-kDa polypeptide (ORF C7L) located in the vaccinia virus HindIII C fragment was shown to be functionally equivalent to previously described host range gene (ORF K1L) spanning the HindIII K/M fragment junction. Either C7L or K1L host range gene is necessary and sufficient by itself to allow replication of vaccinia virus on human cells. Deletion of both C7L and K1L genes from the wild-type vaccinia genome is required to derive a virus deficient for replication on human cells. Further, an ORF encoding a 77-kDa polypeptide derived from cowpox (CP77kDa) and previously shown to allow vaccinia to overcome the restriction for replication on Chinese hamster ovary cells could substitute for the vaccinia host range genes C7L and K1L in permitting replication of the virus on human cells. Additionally, the three unique host range genes C7L, K1L, and CP77kDa were functionally equivalent for vaccinia replication on pig kidney cells, but not on rabbit kidney cells.
    Virology 12/1990; 179(1):276-86. · 3.37 Impact Factor
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    ABSTRACT: Biochemical and genetic analyses have been conducted to determine whether a vaccinia virus open reading frame (orf) with extensive homology to the Saccharomyces cerevisiae DNA ligase gene encodes a functional ligase activity. This orf in HindIII A, designated A50R, is capable of encoding a 552-amino-acid, 63.4-kDa polypeptide. Full-length A50R mRNA produced in vitro directed the synthesis of a polypeptide with an apparent molecular weight of 57 kDa. Significantly, translation reactions programmed with A50R mRNA were capable of ligating a 3-kb Notl restriction fragment into multimers. DNA ligase activity was not detectable when either truncated sense or full-length antisense mRNA was translated in vitro. In extracts prepared from cells infected with wt vaccinia virus, DNA ligase activity was detected as assayed by the formation of a 57 kDa ligase-AMP adduct which was expressed early in the viral replication cycle. In cells infected with a DNA ligase deletion mutant no equivalent AMP-labeled adduct was detected. Relative to wt virus, the DNA ligase deletion mutant exhibited no significant differences in homologous recombination. These results indicate that the vaccinia orf A50R encodes a functional DNA ligase expressed early in infection, but this DNA ligase is nonessential for either recombination or viral replication.
    Virology 12/1990; 179(1):267-75. · 3.37 Impact Factor
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    ABSTRACT: The complete DNA sequence of the genome of vaccinia virus has been determined. The genome consisted of 191,636 bp with a base composition of 66.6% A + T. We have identified 198 "major" protein-coding regions and 65 overlapping "minor" regions, for a total of 263 potential genes. Genes encoded by the virus were located by examination of DNA sequence characteristics and compared with existing vaccinia virus mapping analyses, sequence data, and transcription data. These genes were found to be compactly organized along the genome with relatively few regions of noncoding sequences. Whereas several similarities to proteins of known function were discerned, the function of the majority of proteins encoded by these open reading frames is as yet undetermined.
    Virology 12/1990; 179(1):247-66, 517-63. · 3.37 Impact Factor
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    ABSTRACT: The nucleotide sequence of a 10465 bp HindIII genomic fragment from fowlpox virus (FPV) is presented. Analysis of the nucleotide sequence revealed 10 potential major open reading frames (ORFs). Five of these ORFs are predicted to encode polypeptides with significant homology to hypothetical polypeptides derived from nucleotide sequence analysis of the vaccinia virus (VV) HindIII D region. Interestingly, these homologous ORFs do not occur in the same tandem arrangement in the FPV genome as they do in the VV genome. These results are discussed.
    Journal of General Virology 08/1990; 71 ( Pt 7):1517-24. · 3.13 Impact Factor
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    ABSTRACT: The equine herpesvirus 1 glycoprotein 14 (EHV-1 gp14) gene was cloned, sequenced, and expressed by vaccinia virus recombinants. Recombinant virus vP613 elicited the production of EHV-1-neutralizing antibodies in guinea pigs and was effective in protecting hamsters from subsequent lethal EHV-1 challenge. Coexpression of EHV-1 gp14 in vaccinia virus recombinant vP634 along with EHV-1 gp13 (P. Guo, S. Goebel, S. Davis, M. E. Perkus, B. Languet, P. Desmettre, G. Allen, and E. Paoletti, J. Virol. 63:4189-4198, 1989) greatly enhanced the protective efficacy in the hamster challenge model over that obtained with single recombinants. The inoculum doses (log10) required for protection of 50% of hamsters were 6.1 (EHV-1 gp13), 5.2 (EHV-1 gp14), and less than 3.6 (vaccinia virus recombinant expressing both EHV-1 glycoproteins [gp13 and gp14]).
    Journal of Virology 06/1990; 64(5):2399-406. · 5.08 Impact Factor
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    ABSTRACT: A cDNA copy of the RNA encoding the fusion (F) protein of Newcastle disease virus (NDV) strain Texas, a velogenic strain of NDV, was obtained and the sequence was determined. The 1,792-base-pair sequence encodes a protein of 553 amino acids which has essential features previously established for the F protein of virulent NDV strains. These include the presence of three strongly hydrophobic regions and pairs of dibasic amino acids in the pentapeptide Arg-Arg-Gln-Arg-Arg preceding the putative cleavage site. When inserted into a fowlpox virus vector, a glycosylated protein was expressed and presented on the surface of infected chicken embryo fibroblast cells. The F protein expressed by the recombinant fowlpox virus was cleaved into two polypeptides. When inoculated into susceptible birds by a variety of routes, an immunological response was induced. Ocular or oral administration of the recombinant fowlpox virus gave partial protection, whereas both intramuscular and wing-web routes of inoculation gave complete protection after a single inoculation.
    Journal of Virology 05/1990; 64(4):1441-50. · 5.08 Impact Factor
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    ABSTRACT: The equine herpesvirus 1 (EHV-1) gene encoding glycoprotein 13 (gp13) was cloned into the hemagglutinin (HA) locus of vaccinia virus (Copenhagen strain). Expression of the gp13 gene was driven by the early/late vaccinia virus H6 promoter. Metabolically radiolabeled polypeptides of approximately 47 and 44 kilodaltons and 90 kilodaltons (glycosylated form) were precipitated with both polyclonal and gp13-specific monoclonal antibodies. Presentation of gp13 on the cytoplasmic membrane of cells infected with the recombinant gp13 vaccinia virus was demonstrated by immunofluorescence of unfixed cells. Inoculation of the recombinant gp13 vaccinia virus into guinea pigs induced neutralizing antibodies to both EHV-1 and vaccinia virus. Hamsters vaccinated with the recombinant gp13 vaccinia virus survived a lethal challenge with the hamster-adapted Kentucky strain of EHV-1. These results indicate that expression in vaccinia virus vectors of EHV-1 gp13, the glycoprotein homolog of herpes simplex virus gC-1 and gC-2, pseudorabies virus gIII, and the varicella-zoster virus gpV may provide useful vaccine candidates for equine herpesvirus infections.
    Journal of Virology 11/1989; 63(10):4189-98. · 5.08 Impact Factor
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    ABSTRACT: Theequine herpesvirus 1(EHV-1) geneencoding glycoprotein 13(gpl3) was cloned intothehemagglutinin (HA)locus ofvaccinia virus (Copenhagen strain). Expression ofthegpl3gene was driven bytheearly/late vaccinia virus H6promoter. Metabolically radiolabeled polypeptides ofapproximately 47and44kilodaltons and90kilodaltons (glycosylated form) were precipitated withbothpolyclonal andgpl3-specific monoclonal antibodies. Presentation ofgpl3on thecytoplasmic membraneofcells infected withtherecombinant gpl3 vaccinia virus was demonstrated byimmunofluorescence ofunfixed cells. Inoculation oftherecombinant gpl3 vaccinia virus intoguinea pigsinduced neutralizing antibodies tobothEHV-1andvaccinia virus. Hamsters vaccinated withtherecombinant gpl3vaccinia virus survived a lethal challenge withthehamster-adapted Kentucky strain ofEHV-1.Theseresults indicate thatexpression invaccinia virus vectors ofEHV-1gpl3,the glycoprotein homolog ofherpes simplex virus gC-landgC-2, pseudorabies virusglll, andthevaricella-zoster virus gpVmay provide useful vaccine candidates forequine herpesvirus infections.
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    ABSTRACT: words Text: 2062 words (excluding references and figure legends)

Publication Stats

1k Citations
78.65 Total Impact Points

Institutions

  • 2010
    • New York State Department of Health
      • Wadsworth Center
      New York City, NY, United States
  • 2007–2010
    • Wadsworth Center, NYS Department of Health
      Albany, New York, United States
    • University of California, Davis
      • School of Veterinary Medicine
      Davis, California, United States
  • 2004–2010
    • State University of New York
      New York City, New York, United States
  • 2004–2007
    • University at Albany, The State University of New York
      • Department of Biomedical Sciences
      New York City, NY, United States
  • 1990
    • Albany Medical College
      Albany, New York, United States