Vaccination of Mice with Gonococcal TbpB Expressed In Vivo from Venezuelan Equine Encephalitis Viral Replicon Particles

Thomas, C.; Zhu, Wanling; Dam, Cornelius N. Van; Davis, Nancy; Johnston, Robert E.; Sparling, P. Frederick

doi:10.1128/iai.74.3.1612-1620.2006

Cited by 23 publications

(17 citation statements)

References 48 publications

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“…This is the case for formalininactivated RSV vaccine in infants and more recently FIhMPV vaccine in cotton rats (51). Vaccination with VRPs had been shown to skew toward Th1-type responses (20,37). We therefore evaluated lung histopathology and cytokine gene expression in VRP-vaccinated animals after wild-type hMPV challenge.…”

Section: Discussionmentioning

confidence: 99%

An Alphavirus Replicon-Based Human Metapneumovirus Vaccine Is Immunogenic and Protective in Mice and Cotton Rats

Mok

Tollefson²,

Podsiad³

et al. 2008

J Virol

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Human metapneumovirus (hMPV) is a recently discovered paramyxovirus that causes upper and lower respiratory tract infections in infants, the elderly, and immunocompromised individuals worldwide. Here, we developed Venezuelan equine encephalitis virus replicon particles (VRPs) encoding hMPV fusion (F) or attachment (G) glycoproteins and evaluated the immunogenicity and protective efficacy of these vaccine candidates in mice and cotton rats. VRPs encoding hMPV F protein, when administered intranasally, induced F-specific virus-neutralizing antibodies in serum and immunoglobulin A (IgA) antibodies in secretions at the respiratory mucosa. Challenge virus replication was reduced significantly in both the upper and lower respiratory tracts following intranasal hMPV challenge in these animals. However, vaccination with hMPV G protein VRPs did not induce neutralizing antibodies or protect animals from hMPV challenge. Close examination of the histopathology of the lungs of VRP-MPV F-vaccinated animals following hMPV challenge revealed no enhancement of inflammation or mucus production. Aberrant cytokine gene expression was not detected in these animals. Together, these results represent an important first step toward the use of VRPs encoding hMPV F proteins as a prophylactic vaccine for hMPV.

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Section: Discussionmentioning

confidence: 99%

An Alphavirus Replicon-Based Human Metapneumovirus Vaccine Is Immunogenic and Protective in Mice and Cotton Rats

Mok

Tollefson²,

Podsiad³

et al. 2008

J Virol

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“…TbpA was detected using polyclonal rabbit serum specific for TbpA (17). TbpB was detected on Western blots using a polyclonal anti-TbpB rabbit serum (30) and hTf-HRP (20). Triplicate sets of the hTf-HRP probed blots were scanned and quantitated using NIH ImageJ (31).…”

Section: Methodsmentioning

confidence: 99%

“…Blocked membranes were probed with polyclonal rabbit antiserum against TbpB (30). Goat anti-rabbit IgG conjugated to horseradish peroxidase (HRP) or alkaline phosphatase (AP) was used as the secondary antibody.…”

Section: Methodsmentioning

confidence: 99%

Conserved Regions of Gonococcal TbpB Are Critical for Surface Exposure and Transferrin Iron Utilization

et al. 2013

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The transferrin-binding proteins TbpA and TbpB enable Neisseria gonorrhoeae to obtain iron from human transferrin. The lipoprotein TbpB facilitates, but is not strictly required for, TbpA-mediated iron acquisition. The goal of the current study was to determine the contribution of two conserved regions within TbpB to the function of this protein. Using site-directed mutagenesis, the first mutation we constructed replaced the lipobox (LSAC) of TbpB with a signal I peptidase cleavage site (LAAA), while the second mutation deleted a conserved stretch of glycine residues immediately downstream of the lipobox. We then evaluated the resulting mutants for effects on TbpB expression, surface exposure, and transferrin iron utilization. Western blot analysis and palmitate labeling indicated that the lipobox, but not the glycine-rich motif, is required for lipidation of TbpB and tethering to the outer membrane. TbpB was released into the supernatant by the mutant that produces TbpB LSAC. Neither mutation disrupted the transport of TbpB across the bacterial cell envelope. When these mutant TbpB proteins were produced in a strain expressing a form of TbpA that requires TbpB for iron acquisition, growth on transferrin was either abrogated or dramatically diminished. We conclude that surface tethering of TbpB is required for optimal performance of the transferrin iron acquisition system, while the presence of the polyglycine stretch near the amino terminus of TbpB contributes significantly to transferrin iron transport function. Overall, these results provide important insights into the functional roles of two conserved motifs of TbpB, enhancing our understanding of this critical iron uptake system.

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“…Membranes were developed using Perkin Elmer Chemiluminescence Plus (Perkin Elmer), and reactive bands were detected by exposure to film. To detect TbpB, membranes were blocked with 5% skim milk and incubated with polyclonal anti-TbpB antibody against N. gonorrhoeae strain FA1090 (31,44). Alternatively, to detect Tf-binding by TbpB, membranes were probed with HRP-Tf as described previously (16).…”

Section: Methodsmentioning

confidence: 99%

Identification of Transferrin-Binding Domains in TbpB Expressed by Neisseria gonorrhoeae

DeRocco

Cornelissen

2007

Infect Immun

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The transferrin iron acquisition system of Neisseria gonorrhoeae is necessary for iron uptake from transferrin in the human host and requires the participation of two distinct proteins: TbpA and TbpB. TbpA is a TonB-dependent outer membrane transporter responsible for the transport of iron into the cell. TbpB is a lipid-modified protein, for which a precise role in receptor function has not yet been elucidated. These receptor complex proteins show promise as vaccine candidates; therefore, it is important to identify surface-exposed regions of the proteins required for wild-type functions. In this study we examined TbpB, which has been reported to be surface exposed in its entirety; however, this hypothesis has never been tested experimentally. We placed the hemagglutinin (HA) epitope into TbpB with the dual purpose of examining the surface exposure of particular epitopes as well as their impact on receptor function. Nine insertion mutants were created, placing the epitope downstream of the signal peptidase II cleavage site. We report that the HA epitope is surface accessible in all mutants, indicating that the full-length TbpB is completely surface exposed. By expressing the TbpB-HA fusion proteins in N. gonorrhoeae, we were able to examine the impact of each insertion on the function of TbpB and the transferrin acquisition process. We propose that TbpB is comprised of two transferrin-binding-competent lobes, both of which are critical for efficient iron uptake from human transferrin.

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Vaccination of Mice with Gonococcal TbpB Expressed In Vivo from Venezuelan Equine Encephalitis Viral Replicon Particles

Cited by 23 publications

References 48 publications

An Alphavirus Replicon-Based Human Metapneumovirus Vaccine Is Immunogenic and Protective in Mice and Cotton Rats

An Alphavirus Replicon-Based Human Metapneumovirus Vaccine Is Immunogenic and Protective in Mice and Cotton Rats

Conserved Regions of Gonococcal TbpB Are Critical for Surface Exposure and Transferrin Iron Utilization

Identification of Transferrin-Binding Domains in TbpB Expressed by Neisseria gonorrhoeae

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