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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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Survival curves of mice infected with M. tuberculosis.Survival of parental B6 and I/St and recombinant congenic mice (males) following aerosol challenge with ~500 CFU of M. tuberculosis H37Rv. Recombinant strains B6.I-9.3.19.8, B6.I-9.5.7, B6.I-9.5, B6.I-9.3, B6.I-249.1.15.46 and B6.I-249.1.15 all displayed similar (P > 0.1) intermediate mean survival time (MST) compared to hyper-susceptible I/St and relatively resistant B6 mice (P < 0.0001, log-rank test), which reflects the input of the intra-H2 QTL in susceptibility. All recombinant strains were tested in 3–10 independent experiments (total N = 20–70 animals). Summary of 3–5 experiments is displayed (Kaplan-Meier survival analysis).
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pgen.1005672.g002: Survival curves of mice infected with M. tuberculosis.Survival of parental B6 and I/St and recombinant congenic mice (males) following aerosol challenge with ~500 CFU of M. tuberculosis H37Rv. Recombinant strains B6.I-9.3.19.8, B6.I-9.5.7, B6.I-9.5, B6.I-9.3, B6.I-249.1.15.46 and B6.I-249.1.15 all displayed similar (P > 0.1) intermediate mean survival time (MST) compared to hyper-susceptible I/St and relatively resistant B6 mice (P < 0.0001, log-rank test), which reflects the input of the intra-H2 QTL in susceptibility. All recombinant strains were tested in 3–10 independent experiments (total N = 20–70 animals). Summary of 3–5 experiments is displayed (Kaplan-Meier survival analysis).

Mentions: We transferred genomic regions covering the vicinity of the H2 complex from TB-susceptible parental I/St (H2j) mice onto the B6 (H2b) genetic background in successive backcross generations. Starting with the BC1 (N2) generation, we applied simultaneous selection for the presence of two traits: TB-susceptible phenotype and Chromosome 17 markers of the I/St origin. At the generation N10-11, more than forty B6.I-H2 recombinant congenic strains on the B6 background carrying different, partly-overlapping genomic regions of the extended H2j- haplotype (17 Chr: 8,44–65,34 Mb) were generated. Fig 1 displays the most informative B6.I strains whose pheno- and genotyping allowed us to narrow the region of interest to the interval 33,77–34,34Mb (a total of 0,57 Mb). Mice of all strains which inherited this region from I/St ancestors were significantly more susceptible to infection than those bearing B6 alleles as indicated by survival curves (Fig 2) and the dynamics of cachexia (S1 Fig). Fine mapping within this region was achieved by superposition: the resistant strain B6.I-249.1.16 carries H2j alleles more proximal than the SNPs rs13482956 (17:33, 773331) whereas the strain B6.I-9.3.19.8 is susceptible although all distal genes starting with and including H2-Ea are of B6 origin (Fig 1). Being more susceptible than B6, all recombinant strains carrying the region 33, 77–34, 34Mb inherited from I/St were more resistant than their I/St ancestors, indicating the influence of B6 background genes on survival. This is further supported by the fact that the level of resistance was identical in parental B6 mice and in recombinant mice which inherited the identified H2 segment from B6 (Fig 2). According to http://www.ensembl.org, the identified region contains 36 protein-coding genes, most of which may have important immunological and regulatory functions.


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)

Survival curves of mice infected with M. tuberculosis.Survival of parental B6 and I/St and recombinant congenic mice (males) following aerosol challenge with ~500 CFU of M. tuberculosis H37Rv. Recombinant strains B6.I-9.3.19.8, B6.I-9.5.7, B6.I-9.5, B6.I-9.3, B6.I-249.1.15.46 and B6.I-249.1.15 all displayed similar (P > 0.1) intermediate mean survival time (MST) compared to hyper-susceptible I/St and relatively resistant B6 mice (P < 0.0001, log-rank test), which reflects the input of the intra-H2 QTL in susceptibility. All recombinant strains were tested in 3–10 independent experiments (total N = 20–70 animals). Summary of 3–5 experiments is displayed (Kaplan-Meier survival analysis).
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4664271&req=5

pgen.1005672.g002: Survival curves of mice infected with M. tuberculosis.Survival of parental B6 and I/St and recombinant congenic mice (males) following aerosol challenge with ~500 CFU of M. tuberculosis H37Rv. Recombinant strains B6.I-9.3.19.8, B6.I-9.5.7, B6.I-9.5, B6.I-9.3, B6.I-249.1.15.46 and B6.I-249.1.15 all displayed similar (P > 0.1) intermediate mean survival time (MST) compared to hyper-susceptible I/St and relatively resistant B6 mice (P < 0.0001, log-rank test), which reflects the input of the intra-H2 QTL in susceptibility. All recombinant strains were tested in 3–10 independent experiments (total N = 20–70 animals). Summary of 3–5 experiments is displayed (Kaplan-Meier survival analysis).
Mentions: We transferred genomic regions covering the vicinity of the H2 complex from TB-susceptible parental I/St (H2j) mice onto the B6 (H2b) genetic background in successive backcross generations. Starting with the BC1 (N2) generation, we applied simultaneous selection for the presence of two traits: TB-susceptible phenotype and Chromosome 17 markers of the I/St origin. At the generation N10-11, more than forty B6.I-H2 recombinant congenic strains on the B6 background carrying different, partly-overlapping genomic regions of the extended H2j- haplotype (17 Chr: 8,44–65,34 Mb) were generated. Fig 1 displays the most informative B6.I strains whose pheno- and genotyping allowed us to narrow the region of interest to the interval 33,77–34,34Mb (a total of 0,57 Mb). Mice of all strains which inherited this region from I/St ancestors were significantly more susceptible to infection than those bearing B6 alleles as indicated by survival curves (Fig 2) and the dynamics of cachexia (S1 Fig). Fine mapping within this region was achieved by superposition: the resistant strain B6.I-249.1.16 carries H2j alleles more proximal than the SNPs rs13482956 (17:33, 773331) whereas the strain B6.I-9.3.19.8 is susceptible although all distal genes starting with and including H2-Ea are of B6 origin (Fig 1). Being more susceptible than B6, all recombinant strains carrying the region 33, 77–34, 34Mb inherited from I/St were more resistant than their I/St ancestors, indicating the influence of B6 background genes on survival. This is further supported by the fact that the level of resistance was identical in parental B6 mice and in recombinant mice which inherited the identified H2 segment from B6 (Fig 2). According to http://www.ensembl.org, the identified region contains 36 protein-coding genes, most of which may have important immunological and regulatory functions.

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