Vaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85.

2.50
Hdl Handle:
http://hdl.handle.net/10541/90043
Title:
Vaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85.
Authors:
Malin, Adam S; Huygen, Kris; Content, Jean; Mackett, Mike; Brandt, Lisa; Andersen, Peter; Smith, Steven M; Dockrell, Hazel M
Abstract:
There is increasing evidence to implicate a role for CD8(+) T cells in protective immunity against tuberculosis. Recombinant vaccinia (rVV) expressing Mycobacterium tuberculosis (MTB) proteins can be used both as tools to dissect CD8(+) T-cell responses and, in attenuated form, as candidate vaccines capable of inducing a balanced CD4(+)/CD8(+) T-cell response. A panel of rVV was constructed to express four immunodominant secreted proteins of MTB: 85A, 85B and 85C and ESAT-6. A parallel group of rVV was constructed to include the heterologous eukaryotic tissue plasminogen activator (tPA) signal sequence to assess if this would enhance expression and immunogenicity. Clear expression was obtained for 85A, 85B and ESAT-6 and the addition of tPA resulted in N-glycosylation and a 4-10-fold increase in expression. Female C57BL/6 mice were immunised using the rVV-Ag85 constructs, and interleukin-2 and gamma-interferon were assayed using a co-culture of immune splenocytes and recall antigen. There was a marked increase in cytokine production in mice immunised with the tPA-containing constructs. We report the first data demonstrating enhanced immunogenicity of rVV using a tPA signal sequence, which has significant implications for future vaccine design.
Affiliation:
Immunology Unit, Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, WC1E 7HT, London, UK. adam.malin@bigfoot.com
Citation:
Vaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85. 2000, 2 (14):1677-85 Microbes Infect.
Journal:
Microbes and Infection
Issue Date:
Nov-2000
URI:
http://hdl.handle.net/10541/90043
DOI:
10.1016/S1286-4579(00)01323-X
PubMed ID:
11137041
Type:
Article
Language:
en
ISSN:
1286-4579
Appears in Collections:
All Paterson Institute for Cancer Research

Full metadata record

DC FieldValue Language
dc.contributor.authorMalin, Adam Sen
dc.contributor.authorHuygen, Krisen
dc.contributor.authorContent, Jeanen
dc.contributor.authorMackett, Mikeen
dc.contributor.authorBrandt, Lisaen
dc.contributor.authorAndersen, Peteren
dc.contributor.authorSmith, Steven Men
dc.contributor.authorDockrell, Hazel Men
dc.date.accessioned2010-01-19T17:25:05Z-
dc.date.available2010-01-19T17:25:05Z-
dc.date.issued2000-11-
dc.identifier.citationVaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85. 2000, 2 (14):1677-85 Microbes Infect.en
dc.identifier.issn1286-4579-
dc.identifier.pmid11137041-
dc.identifier.doi10.1016/S1286-4579(00)01323-X-
dc.identifier.urihttp://hdl.handle.net/10541/90043-
dc.description.abstractThere is increasing evidence to implicate a role for CD8(+) T cells in protective immunity against tuberculosis. Recombinant vaccinia (rVV) expressing Mycobacterium tuberculosis (MTB) proteins can be used both as tools to dissect CD8(+) T-cell responses and, in attenuated form, as candidate vaccines capable of inducing a balanced CD4(+)/CD8(+) T-cell response. A panel of rVV was constructed to express four immunodominant secreted proteins of MTB: 85A, 85B and 85C and ESAT-6. A parallel group of rVV was constructed to include the heterologous eukaryotic tissue plasminogen activator (tPA) signal sequence to assess if this would enhance expression and immunogenicity. Clear expression was obtained for 85A, 85B and ESAT-6 and the addition of tPA resulted in N-glycosylation and a 4-10-fold increase in expression. Female C57BL/6 mice were immunised using the rVV-Ag85 constructs, and interleukin-2 and gamma-interferon were assayed using a co-culture of immune splenocytes and recall antigen. There was a marked increase in cytokine production in mice immunised with the tPA-containing constructs. We report the first data demonstrating enhanced immunogenicity of rVV using a tPA signal sequence, which has significant implications for future vaccine design.en
dc.language.isoenen
dc.subject.meshAcyltransferases-
dc.subject.meshAnimals-
dc.subject.meshAntigens, Bacterial-
dc.subject.meshBacterial Proteins-
dc.subject.meshBacterial Vaccines-
dc.subject.meshBase Sequence-
dc.subject.meshCells, Cultured-
dc.subject.meshCloning, Molecular-
dc.subject.meshCoculture Techniques-
dc.subject.meshFemale-
dc.subject.meshGenetic Vectors-
dc.subject.meshGlycosylation-
dc.subject.meshInterferon-gamma-
dc.subject.meshInterleukin-2-
dc.subject.meshMice-
dc.subject.meshMice, Inbred C57BL-
dc.subject.meshMolecular Sequence Data-
dc.subject.meshMycobacterium tuberculosis-
dc.subject.meshProtein Folding-
dc.subject.meshSignal Recognition Particle-
dc.subject.meshTissue Plasminogen Activator-
dc.subject.meshVaccinia virus-
dc.titleVaccinia expression of Mycobacterium tuberculosis-secreted proteins: tissue plasminogen activator signal sequence enhances expression and immunogenicity of M. tuberculosis Ag85.en
dc.typeArticleen
dc.contributor.departmentImmunology Unit, Department of Infectious and Tropical Diseases, London School of Hygiene & Tropical Medicine, WC1E 7HT, London, UK. adam.malin@bigfoot.comen
dc.identifier.journalMicrobes and Infectionen
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