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The QTL within the H2 Complex Involved in the Control of Tuberculosis Infection in Mice Is the Classical Class II H2-Ab1 Gene.

Logunova N, Korotetskaya M, Polshakov V, Apt A - PLoS Genet. (2015)

Bottom Line: Cloning and sequencing of the H2j allelic variants of these genes demonstrated profound polymorphic variations compare to the H2b haplotype.These variations were sufficient to produce different TB-relevant phenotypes: the more susceptible B6.I-249.1.15.100 strain demonstrated shorter survival time, more rapid body weight loss, higher mycobacterial loads in the lungs and more severe lung histopathology compared to the more resistant B6.I-249.1.15.139 strain.CD4+ T cells recognized mycobacterial antigens exclusively in the context of the H2-A Class II molecule, and the level of IFN-γ-producing CD4+ T cells in the lungs was significantly higher in the resistant strain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Immunogenetics, Central Institute for Tuberculosis, Moscow, Russia.

ABSTRACT
The level of susceptibility to tuberculosis (TB) infection depends upon allelic variations in numerous interacting genes. In our mouse model system, the whole-genome quantitative trait loci (QTLs) scan revealed three QTLs involved in TB control on chromosomes 3, 9, and in the vicinity of the H2 complex on chromosome 17. For the present study, we have established a panel of new congenic, MHC-recombinant mouse strains bearing differential small segments of chromosome 17 transferred from the TB-susceptible I/St (H2j) strain onto the genetic background of TB-resistant C57BL/6 (B6) mice (H2b). This allowed narrowing the QTL interval to 17Ch: 33, 77-34, 34 Mb, containing 36 protein-encoding genes. Cloning and sequencing of the H2j allelic variants of these genes demonstrated profound polymorphic variations compare to the H2b haplotype. In two recombinant strains, B6.I-249.1.15.100 and B6.I-249.1.15.139, recombination breakpoints occurred in different sites of the H2-Aβ 1 gene (beta-chain of the Class II heterodimer H2-A), providing polymorphic variations in the domain β1 of the Aβ-chain. These variations were sufficient to produce different TB-relevant phenotypes: the more susceptible B6.I-249.1.15.100 strain demonstrated shorter survival time, more rapid body weight loss, higher mycobacterial loads in the lungs and more severe lung histopathology compared to the more resistant B6.I-249.1.15.139 strain. CD4+ T cells recognized mycobacterial antigens exclusively in the context of the H2-A Class II molecule, and the level of IFN-γ-producing CD4+ T cells in the lungs was significantly higher in the resistant strain. Thus, we directly demonstrated for the first time that the classical H2- Ab1 Class II gene is involved in TB control. Molecular modeling of the H2-Aj product predicts that amino acid (AA) substitutions in the Aβ-chain modify the motif of the peptide-MHC binding groove. Moreover, unique AA substitutions in both α- and β-chains of the H2-Aj molecule might affect its interactions with the T-cell receptor (TCR).

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CD4+ T cells recognize mycobacterial antigens in the context of H2-A molecule.Mycobacterial antigens were presented by APC expressing different gene combinations and alleles of Class II molecules (see legend) to: (A) polyclonal I/St T cell line, or to highly purified immune lymph node CD4+ T cells (pooled from 2–3 mice) from I/St (B), B6.I-100 (C) and B6.I.139 mice (D). Representative data from one out of two similar independent experiments are presented, results are expressed as mean ± SEM of triplicate cultures. Y-axis: Δ cpm (counts per minute) = mean cpm of antigen-stimulated wells—the mean cpm of non-stimulated wells. Stimulation index (SI) mean cpm of antigen-stimulated wells/ mean cpm of non-stimulated wells. (E)–CD4+ T-cells from (B6 x B6.I-9.3.19.8) F1 mice stimulate bacteriostatic activity of B6 and F1, but not of B6.I-9.3.19.8 macrophages (mph). Results are present as [3H]-uracil uptake from one out of two experiments provided similar results (CPM ± SEM for triplicates, P < 0.001, ANOVA). See Materials and Methods for details.
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pgen.1005672.g006: CD4+ T cells recognize mycobacterial antigens in the context of H2-A molecule.Mycobacterial antigens were presented by APC expressing different gene combinations and alleles of Class II molecules (see legend) to: (A) polyclonal I/St T cell line, or to highly purified immune lymph node CD4+ T cells (pooled from 2–3 mice) from I/St (B), B6.I-100 (C) and B6.I.139 mice (D). Representative data from one out of two similar independent experiments are presented, results are expressed as mean ± SEM of triplicate cultures. Y-axis: Δ cpm (counts per minute) = mean cpm of antigen-stimulated wells—the mean cpm of non-stimulated wells. Stimulation index (SI) mean cpm of antigen-stimulated wells/ mean cpm of non-stimulated wells. (E)–CD4+ T-cells from (B6 x B6.I-9.3.19.8) F1 mice stimulate bacteriostatic activity of B6 and F1, but not of B6.I-9.3.19.8 macrophages (mph). Results are present as [3H]-uracil uptake from one out of two experiments provided similar results (CPM ± SEM for triplicates, P < 0.001, ANOVA). See Materials and Methods for details.

Mentions: A mycobacteria-specific CD4+ T cell line derived from I/St mice [49] readily proliferated in the presence of mycobacterial antigens if the APC were derived from mice expressing the H2-Aj molecule, even if the H2-E molecule was not expressed (Fig 6A). Moreover, the presence or absence of H2-E did not change the level of response, suggesting that the H2-A molecule presents mycobacterial antigens to the vast majority of T-cell clones. To prove that the H2-E-recognizing T-cell clones have not been lost due to repeated stimulation during T-cell line development, we repeated the experiment with highly purified CD4+ T cells from TB-immune lymph nodes of I/St mice and obtained similar results (Fig 6B).


The QTL within the H2 Complex Involved in the Control of Tuberculosis Infection in Mice Is the Classical Class II H2-Ab1 Gene.

Logunova N, Korotetskaya M, Polshakov V, Apt A - PLoS Genet. (2015)

CD4+ T cells recognize mycobacterial antigens in the context of H2-A molecule.Mycobacterial antigens were presented by APC expressing different gene combinations and alleles of Class II molecules (see legend) to: (A) polyclonal I/St T cell line, or to highly purified immune lymph node CD4+ T cells (pooled from 2–3 mice) from I/St (B), B6.I-100 (C) and B6.I.139 mice (D). Representative data from one out of two similar independent experiments are presented, results are expressed as mean ± SEM of triplicate cultures. Y-axis: Δ cpm (counts per minute) = mean cpm of antigen-stimulated wells—the mean cpm of non-stimulated wells. Stimulation index (SI) mean cpm of antigen-stimulated wells/ mean cpm of non-stimulated wells. (E)–CD4+ T-cells from (B6 x B6.I-9.3.19.8) F1 mice stimulate bacteriostatic activity of B6 and F1, but not of B6.I-9.3.19.8 macrophages (mph). Results are present as [3H]-uracil uptake from one out of two experiments provided similar results (CPM ± SEM for triplicates, P < 0.001, ANOVA). See Materials and Methods for details.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4664271&req=5

pgen.1005672.g006: CD4+ T cells recognize mycobacterial antigens in the context of H2-A molecule.Mycobacterial antigens were presented by APC expressing different gene combinations and alleles of Class II molecules (see legend) to: (A) polyclonal I/St T cell line, or to highly purified immune lymph node CD4+ T cells (pooled from 2–3 mice) from I/St (B), B6.I-100 (C) and B6.I.139 mice (D). Representative data from one out of two similar independent experiments are presented, results are expressed as mean ± SEM of triplicate cultures. Y-axis: Δ cpm (counts per minute) = mean cpm of antigen-stimulated wells—the mean cpm of non-stimulated wells. Stimulation index (SI) mean cpm of antigen-stimulated wells/ mean cpm of non-stimulated wells. (E)–CD4+ T-cells from (B6 x B6.I-9.3.19.8) F1 mice stimulate bacteriostatic activity of B6 and F1, but not of B6.I-9.3.19.8 macrophages (mph). Results are present as [3H]-uracil uptake from one out of two experiments provided similar results (CPM ± SEM for triplicates, P < 0.001, ANOVA). See Materials and Methods for details.
Mentions: A mycobacteria-specific CD4+ T cell line derived from I/St mice [49] readily proliferated in the presence of mycobacterial antigens if the APC were derived from mice expressing the H2-Aj molecule, even if the H2-E molecule was not expressed (Fig 6A). Moreover, the presence or absence of H2-E did not change the level of response, suggesting that the H2-A molecule presents mycobacterial antigens to the vast majority of T-cell clones. To prove that the H2-E-recognizing T-cell clones have not been lost due to repeated stimulation during T-cell line development, we repeated the experiment with highly purified CD4+ T cells from TB-immune lymph nodes of I/St mice and obtained similar results (Fig 6B).

Bottom Line: Cloning and sequencing of the H2j allelic variants of these genes demonstrated profound polymorphic variations compare to the H2b haplotype.These variations were sufficient to produce different TB-relevant phenotypes: the more susceptible B6.I-249.1.15.100 strain demonstrated shorter survival time, more rapid body weight loss, higher mycobacterial loads in the lungs and more severe lung histopathology compared to the more resistant B6.I-249.1.15.139 strain.CD4+ T cells recognized mycobacterial antigens exclusively in the context of the H2-A Class II molecule, and the level of IFN-γ-producing CD4+ T cells in the lungs was significantly higher in the resistant strain.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Immunogenetics, Central Institute for Tuberculosis, Moscow, Russia.

ABSTRACT
The level of susceptibility to tuberculosis (TB) infection depends upon allelic variations in numerous interacting genes. In our mouse model system, the whole-genome quantitative trait loci (QTLs) scan revealed three QTLs involved in TB control on chromosomes 3, 9, and in the vicinity of the H2 complex on chromosome 17. For the present study, we have established a panel of new congenic, MHC-recombinant mouse strains bearing differential small segments of chromosome 17 transferred from the TB-susceptible I/St (H2j) strain onto the genetic background of TB-resistant C57BL/6 (B6) mice (H2b). This allowed narrowing the QTL interval to 17Ch: 33, 77-34, 34 Mb, containing 36 protein-encoding genes. Cloning and sequencing of the H2j allelic variants of these genes demonstrated profound polymorphic variations compare to the H2b haplotype. In two recombinant strains, B6.I-249.1.15.100 and B6.I-249.1.15.139, recombination breakpoints occurred in different sites of the H2-Aβ 1 gene (beta-chain of the Class II heterodimer H2-A), providing polymorphic variations in the domain β1 of the Aβ-chain. These variations were sufficient to produce different TB-relevant phenotypes: the more susceptible B6.I-249.1.15.100 strain demonstrated shorter survival time, more rapid body weight loss, higher mycobacterial loads in the lungs and more severe lung histopathology compared to the more resistant B6.I-249.1.15.139 strain. CD4+ T cells recognized mycobacterial antigens exclusively in the context of the H2-A Class II molecule, and the level of IFN-γ-producing CD4+ T cells in the lungs was significantly higher in the resistant strain. Thus, we directly demonstrated for the first time that the classical H2- Ab1 Class II gene is involved in TB control. Molecular modeling of the H2-Aj product predicts that amino acid (AA) substitutions in the Aβ-chain modify the motif of the peptide-MHC binding groove. Moreover, unique AA substitutions in both α- and β-chains of the H2-Aj molecule might affect its interactions with the T-cell receptor (TCR).

Show MeSH
Related in: MedlinePlus