Immunostimulatory Activity of Recombinant Mycobacterium bovis BCG That Secretes Major Membrane Protein II of Mycobacterium leprae

ABSTRACT We previously demonstrated that major membrane protein II (MMP-II) is one of the immunodominant antigens (Ags) of Mycobacterium leprae capable of activating T cells through Toll-like receptor 2. Based on the observation that Mycobacterium bovis BCG secreting a 30-kDa protein offered better protection against tuberculosis, we constructed a recombinant BCG strain (BCG-SM) that secretes MMP-II to improve the potency of BCG against leprosy. The secreted MMP-II protein from BCG-SM stimulated monocyte-derived dendritic cells (DC) to produce interleukin-12. DC infected with BCG-SM expressed MMP-II on their surfaces, and MMP-II expression was suppressed by the pretreatment of DC with chloroquine. These results indicated that secreted MMP-II was processed by DC for higher expression levels on their surfaces. In addition, BCG-SM phenotypically activated DC and induced higher expression levels of major histocompatibility complex, CD86, and CD83 Ags on DC than did vector control BCG (BCG-pMV). The DC infected with BCG-SM more efficiently stimulated naïve and memory CD4+ T cells and memory CD8+ T cells to produce gamma interferon than did those infected with BCG-pMV. However, naïve CD8+ T cells were significantly activated only when they were stimulated with BCG-SM-infected DC. When CD8+ T cells were cocultured with BCG-SM-infected DC, the proportion of perforin-producing T cells was significantly higher than that in cells cocultured with BCG-pMV-infected DC. Moreover, MMP-II-specific memory T cells were more efficiently produced in mice inoculated with BCG-SM than in mice inoculated with BCG-pMV. Taken together, these results indicate that BCG capable of secreting the immunodominant Ag is more potent in the stimulation of T cells.

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Immunostimulatory Activity of Major Membrane Protein II fromMycobacterium tuberculosis

Clinical and Vaccine Immunology ◽

10.1128/cvi.00459-10 ◽

2010 ◽

Vol 18 (2) ◽

pp. 235-242 ◽

Cited By ~ 7

Author(s):

Yumiko Tsukamoto ◽

Masumi Endoh ◽

Tetsu Mukai ◽

Yumi Maeda ◽

Toshiki Tamura ◽

...

Keyword(s):

T Cells ◽

Membrane Protein ◽

Recombinant Proteins ◽

Interleukin 12 ◽

Toll Like Receptor ◽

Healthy Individuals ◽

Immunostimulatory Activity ◽

Toll Like Receptor 2 ◽

Protein Memory ◽

Memory Type

ABSTRACTPreviously, we observed that both major membrane protein II ofMycobacterium leprae(MMP-ML) and its fusion withM. bovisBCG (BCG)-derived heat shock protein 70 (HSP70) (Fusion-ML) are immunogenic and that recombinant BCG secreting either of these proteins effectively inhibits the multiplication ofM. lepraein mice. Here, we purifiedM. tuberculosis-derived major membrane protein II (MMP-MTB) and its fusion with HSP70 (Fusion-MTB) in a lipopolysaccharide-free condition and evaluated their immunostimulatory abilities. Both MMP-MTB and Fusion-MTB activated monocyte-derived dendritic cells (DC) in terms of phenotype and interleukin-12 (IL-12) production, but Fusion-MTB more efficiently activated them than MMP-MTB did. The IL-12 production was a consequence of the ligation of those recombinant proteins with Toll-like receptor 2. TheM. tuberculosis-derived andM. leprae-derived recombinant proteins activated naïve T cells of both CD4 and CD8 subsets, butM. tuberculosis-derived proteins were superior toM. leprae-derived proteins and fusion proteins were superior to MMP, regardless of the origin of the protein. Memory-type CD4+T cells obtained from BCG-vaccinated healthy individuals seem to be primed with MMP-MTB by the vaccination, and bothM. tuberculosis-derived recombinant proteins produced perforin-producing CD8+T cells from memory-type CD8+T cells. Further, infection of DC and macrophages withM. tuberculosisH37Ra and H37Rv induced the expression of MMP on their surface. These results indicate thatM. tuberculosis-derived MMP, as a sole protein or as part of a fusion protein, may be useful for developing new vaccinating agents against tuberculosis.

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Identification of an Immunomodulating Agent from Mycobacterium leprae

Infection and Immunity ◽

10.1128/iai.73.5.2744-2750.2005 ◽

2005 ◽

Vol 73 (5) ◽

pp. 2744-2750 ◽

Cited By ~ 32

Author(s):

Yumi Maeda ◽

Tetsu Mukai ◽

John Spencer ◽

Masahiko Makino

Keyword(s):

Mycobacterium Leprae ◽

Surface Expression ◽

Interleukin 12 ◽

Toll Like Receptor ◽

Healthy Individuals ◽

Reactive Protein ◽

Histocompatibility Complex ◽

Toll Like Receptor 2 ◽

Antigen Presenting ◽

Dc Maturation

ABSTRACT A search for an immunomodulating agent from mycobacteria was carried out using Mycobacterium leprae. The antigenicity of each fraction of the bacterial membrane, which contains the most antigenic components of M. leprae, was assessed by using sera from paucibacillary leprosy. N-terminal sequencing of the serum-reactive protein and functional assessment of the membrane fractions using monocyte-derived dendritic cells (DCs) identified major membrane protein II (MMP-II) as one of the efficient T-cell-activating candidates. Purified MMP-II stimulated DCs from healthy individuals to produce interleukin-12 p70 and up-regulated the surface expression of major histocompatibility complex class I and II, CD86, and CD83 molecules. Also, there was an increase in the percentage of CD83+ cells in the DC population. Furthermore, MMP-II-pulsed DCs expressed their derivatives on their surfaces. Using Toll-like receptor 2 (TLR-2)-dependent receptor constructs, we found that TLR-2 signaling was involved in DC maturation induced by MMP-II. Taken together, MMP-II can be recognized as an immunomodulating protein in terms of activation of antigen-presenting cells and innate immunity.

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Inhibition of the Multiplication of Mycobacterium leprae by Vaccination with a Recombinant M. bovis BCG Strain That Secretes Major Membrane Protein II in Mice

Clinical and Vaccine Immunology ◽

10.1128/cvi.00203-09 ◽

2009 ◽

Vol 16 (10) ◽

pp. 1399-1404 ◽

Cited By ~ 5

Author(s):

Y. Maeda ◽

T. Tamura ◽

M. Matsuoka ◽

M. Makino

Keyword(s):

T Cells ◽

Membrane Protein ◽

Vector Control ◽

Mycobacterium Bovis ◽

Membrane Fraction ◽

Vaccine Candidate ◽

Mycobacterium Leprae ◽

Cytosolic Fraction ◽

Ifn Γ

ABSTRACT The ability of a recombinant Mycobacterium bovis BCG strain that secretes major membrane protein II (MMP-II) of Mycobacterium leprae (BCG-SM) to confer protection against leprosy was evaluated by use of a mouse footpad model. C57BL/6J mice intradermally inoculated with BCG-SM produced splenic T cells which secreted significant amounts of gamma interferon (IFN-γ) in response to either the recombinant MMP-II, the M. leprae-derived membrane fraction, or the BCG-derived cytosolic fraction in vitro more efficiently than those from the mice infected with the vector control BCG strain (BCG-pMV, a BCG strain containing pMV-261). A higher percentage of CD8+ T cells obtained from BCG-SM-inoculated mice than those obtained from BCG-pMV-inoculated mice produced intracellular IFN-γ on restimulation with the M. leprae antigens. BCG-SM inhibited the multiplication of M. leprae in the footpads of C57BL/6J mice more efficiently than BCG-pMV. These results indicate that a BCG strain that secretes MMP-II could be a better vaccine candidate for leprosy.

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WDFY4 is required for cross-presentation in response to viral and tumor antigens

Science ◽

10.1126/science.aat5030 ◽

2018 ◽

Vol 362 (6415) ◽

pp. 694-699 ◽

Cited By ~ 69

Author(s):

Derek J. Theisen ◽

Jesse T. Davidson ◽

Carlos G. Briseño ◽

Marco Gargaro ◽

Elvin J. Lauron ◽

...

Keyword(s):

T Cells ◽

Dendritic Cells ◽

Tumor Antigens ◽

Critical Role ◽

Tumor Rejection ◽

Interleukin 12 ◽

Cross Presentation ◽

Histocompatibility Complex ◽

Crispr Screen

During the process of cross-presentation, viral or tumor-derived antigens are presented to CD8+ T cells by Batf3-dependent CD8α+/XCR1+ classical dendritic cells (cDC1s). We designed a functional CRISPR screen for previously unknown regulators of cross-presentation, and identified the BEACH domain–containing protein WDFY4 as essential for cross-presentation of cell-associated antigens by cDC1s in mice. However, WDFY4 was not required for major histocompatibility complex class II presentation, nor for cross-presentation by monocyte-derived dendritic cells. In contrast to Batf3–/– mice, Wdfy4–/– mice displayed normal lymphoid and nonlymphoid cDC1 populations that produce interleukin-12 and protect against Toxoplasma gondii infection. However, similar to Batf3–/– mice, Wdfy4–/– mice failed to prime virus-specific CD8+ T cells in vivo or induce tumor rejection, revealing a critical role for cross-presentation in antiviral and antitumor immunity.

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Distinct Roles for MyD88 and Toll-Like Receptor 2 during Leishmania braziliensis Infection in Mice

Infection and Immunity ◽

10.1128/iai.00154-09 ◽

2009 ◽

Vol 77 (7) ◽

pp. 2948-2956 ◽

Cited By ~ 59

Author(s):

Diego A. Vargas-Inchaustegui ◽

Wendy Tai ◽

Lijun Xin ◽

Alison E. Hogg ◽

David B. Corry ◽

...

Keyword(s):

T Cells ◽

Protective Immunity ◽

Interleukin 12 ◽

Leishmania Braziliensis ◽

Toll Like Receptor ◽

Enhanced Resistance ◽

Toll Like Receptor 2 ◽

Resistance To Infection

ABSTRACT We have previously reported that Leishmania braziliensis infection can activate murine dendritic cells (DCs) and upregulate signaling pathways that are essential for the initiation of innate immunity. However, it remains unclear whether Toll-like receptors (TLRs) are involved in L. braziliensis-mediated DC activation. To address this issue, we generated bone marrow-derived DCs from MyD88−/− and TLR2−/− mice and examined their responsiveness to parasite infection. While wild-type DCs were efficiently activated to produce cytokines and prime naïve CD4+ T cells, L. braziliensis-infected MyD88−/− DCs exhibited less activation and decreased production of interleukin-12 (IL-12) p40. Furthermore, MyD88−/− mice were more susceptible to infection in that they developed larger and prolonged lesions compared to those in control mice. In sharp contrast, the lack of TLR2 resulted in an enhanced DC activation and increased IL-12 p40 production after infection. As such, L. braziliensis-infected TLR2−/− DCs were more competent in priming naïve CD4+ T cells in vitro than were their controls, findings which correlated with an increased gamma interferon production in vivo and enhanced resistance to infection. Our results suggest that while MyD88 is indispensable for the generation of protective immunity to L. braziliensis, TLR2 seems to have a regulatory role during infection.

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Mycobacterium tuberculosis Lipomannan Induces Apoptosis and Interleukin-12 Production in Macrophages

Infection and Immunity ◽

10.1128/iai.72.4.2067-2074.2004 ◽

2004 ◽

Vol 72 (4) ◽

pp. 2067-2074 ◽

Cited By ~ 102

Author(s):

D. N. Dao ◽

L. Kremer ◽

Y. Guérardel ◽

A. Molano ◽

W. R. Jacobs ◽

...

Keyword(s):

Cell Wall ◽

Mycobacterium Tuberculosis ◽

Function Analysis ◽

Specific Interaction ◽

Interleukin 12 ◽

Mycobacterial Species ◽

Immunostimulatory Activity ◽

Mycobacterium Chelonae ◽

Toll Like Receptor 2 ◽

Mycobacterial Cell Wall

ABSTRACT The mycobacterial cell wall component lipoarabinomannan (LAM) has been described as a virulence factor of Mycobacterium tuberculosis, and modification of the terminal arabinan residues of this compound with mannose caps (producing mannosyl-capped LAM [ManLAM]) in M. tuberculosis or with phosphoinositol caps (producing phosphoinositol-capped LAM [PILAM]) in Mycobacterium smegmatis has been implicated in various functions associated with these lipoglycans. A structure-function analysis was performed by using LAMs and their biosynthetic precursor lipomannans (LMs) isolated from different mycobacterial species on the basis of their capacity to induce the production of interleukin-12 (IL-12) and/or apoptosis of macrophage cell lines. Independent of the mycobacterial species, ManLAMs did not induce IL-12 gene expression or apoptosis of macrophages, whereas PILAMs induced IL-12 secretion and apoptosis. Interestingly, uncapped LAM purified from Mycobacterium chelonae did not induce IL-12 secretion or apoptosis. Furthermore, LMs, independent of their mycobacterial origins, were potent inducers of IL-12 and apoptosis. The precursor of LM, phosphatidyl-myo-inositol dimannoside, had no activity, suggesting that the mannan core of LM was required for the activity of LM. The specific interaction of LM with Toll-like receptor 2 (TLR-2) but not with TLR-4 suggested that these responses were mediated via the TLR-2 signaling pathway. Our experiments revealed an important immunostimulatory activity of the biosynthetic LAM precursor LM. The ratio of LAM to LM in the cell wall of mycobacteria may be an important determinant of virulence, and enzymes that modify LM could provide targets for development of antituberculosis drugs and for derivation of attenuated strains of M. tuberculosis.

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CD4 memory T cells survive and proliferate but fail to differentiate in the absence of CD40

Journal of Experimental Medicine ◽

10.1084/jem.20050711 ◽

2006 ◽

Vol 203 (4) ◽

pp. 897-906 ◽

Cited By ~ 37

Author(s):

Megan MacLeod ◽

Mark J. Kwakkenbos ◽

Alison Crawford ◽

Sheila Brown ◽

Brigitta Stockinger ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Memory T Cells ◽

Cell Receptor ◽

Memory Cells ◽

Cell Stage ◽

Effector Cells ◽

Cell Responses ◽

Histocompatibility Complex ◽

Specific Memory

Secondary T cell responses are enhanced because of an expansion in numbers of antigen-specific (memory) cells. Using major histocompatibility complex class II tetramers we have tracked peptide-specific endogenous (non–T cell receptor transgenic) CD4 memory T cells in normal and in costimulation-deficient mice. CD4 memory T cells were detectable after immunization for more than 200 days, although decay was apparent. Memory cells generated in CD40 knockout mice by immunization with peptide-pulsed wild-type dendritic cells survived in the absence of CD40 and proliferated when boosted with peptide (plus adjuvant) in a CD40-independent fashion. However, differentiation of the memory cells into cytokine-producing effector cells did not occur in the absence of CD40. The data indicate that memory cells can be generated without passing through the effector cell stage.

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