Molecular basis of pathogenicity in vaccinia virus.
Item
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Title
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Molecular basis of pathogenicity in vaccinia virus.
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Identifier
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AAI9405509
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identifier
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9405509
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Creator
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Chang, Pi-Yun.
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Contributor
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Adviser: Beatriz G.-T. Pogo
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Date
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1993
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Language
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English
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Publisher
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City University of New York.
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Subject
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Biology, Microbiology | Biology, General
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Abstract
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Mutant Z-19 has been isolated from Friend erythroleukemia cells persistently infected with vaccinia virus and characterized. It had 20-21 kb deletion at the left end, including HindIII C and part of HindIII N fragments, and rearrangements at the right end of viral N genome. Since mutant Z-29 provides a null virulence background, it is a good model system to study pathogenicity. Furthermore, it was previously reported that in Jurkat cells persistently infected with vaccinia virus, there is stimulation of cytokine synthesis (IL-2, IL-2 receptor-a, and IL-6) and increase in the transactivation of HIV-1LTR transcription.;The aim of this thesis was to study the molecular pathogenicity of vaccinia virus and to investigate virus-host interaction in Jurkat cells persistently infected with vaccinia virus. Three aspects were explored. (1) To determine if the virulent phenotype can be recovered by introduction of the terminal HindIII C fragment into the attenuated mutant Z-19, rescue experiments were performed, and recombinant viruses were isolated and characterized. The results showed that recovery of virulence in mice was correlated with the presence and expression of two genes: vaccinia growth factor and vaccinia complement-binding protein, both located at the left terminus. (2) To establish a model system to identify which gene(s) is/are responsible for virulence in the IHD-W strain, a shuttle vector was constructed to reintroduce genes into the mutant Z-19. The shuttle vector contained the fusion fragment K{dollar}\sp\prime{dollar} from mutant Z-19, which comprised the end terminal HindIII C and part of the N fragment. The vector also contained the {dollar}\beta{dollar}-galactosidase cassette which included the LacZ gene under the vaccinia promoter P11 for screening purposes. The vaccinia growth factor and vaccinia complement-binding protein genes were inserted back into mutant Z-19. The results showed that these genes were expressed in cells infected with the recombinant viruses and that they became virulent. However, they did not attain the virulence level of the wild-type, suggesting that expression of other genes present in fragment HindIII C may be required. (3) To study the molecular mechanism by which Jurkat cells persistently infected with vaccinia virus transactivated the LTR of HIV-1 virus, gel mobility shift assays were performed using labeled LTR or synthetic oligonucleotides and nuclear extracts derived from activated or infected Jurkat cells to define which sequences in the LTR were involved. Results indicated that two elements: NFAT-1 and NF-kB, were able to interact with nuclear extracts derived from Jurkat cells persistently infected with vaccinia virus, suggesting that these elements may play a role in the transactivation of HIV-LTR. (Abstract shortened by UMI.).
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Type
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dissertation
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Source
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PQT Legacy CUNY.xlsx
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degree
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Ph.D.
