Limits...
Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire.

Trad A, Tanasa RI, Lange H, Zemlin M, Schroeder HW, Lemke H - Front Immunol (2014)

Bottom Line: We chose this antigen because studies of the humoral immune response to the hapten phOx were instrumental in the development of the current theoretical framework on which our understanding of the forces driving TD responses is based.Compared to WT, TD-induced humoral IgM as well as IgG antibody production in the D-altered ΔD-DμFS and ΔD-iD strains were significantly reduced.These changes in clonal selection and expansion, as well as CSR indicate that the genetic constitution of the DH locus, which has been selected by evolution, can strongly influence the functional outcome of a TD humoral response.

View Article: PubMed Central - PubMed

Affiliation: Biochemical Institute of the Medical Faculty of the Christian-Albrechts-University , Kiel , Germany.

ABSTRACT
The diversity of the third complementarity determining region of the IgH chain is constrained by natural selection of immunoglobulin diversity (DH) sequence. To test the functional significance of this constraint in the context of thymus-dependent (TD) immune responses, we immunized BALB/c mice with WT or altered DH sequence with 2-phenyloxazolone-coupled chicken serum albumin (phOx-CSA). We chose this antigen because studies of the humoral immune response to the hapten phOx were instrumental in the development of the current theoretical framework on which our understanding of the forces driving TD responses is based. To allow direct comparison, we used the classic approach of generating monoclonal Ab (mAb) from various stages of the immune response to phOx to assess the effect of changing the sequence of the DH on clonal expansion, class switching, and affinity maturation, which are hallmarks of TD responses. Compared to WT, TD-induced humoral IgM as well as IgG antibody production in the D-altered ΔD-DμFS and ΔD-iD strains were significantly reduced. An increased prevalence of IgM-producing hybridomas from late primary, secondary, and tertiary memory responses suggested either impaired class switch recombination (CSR) or impaired clonal expansion of class switched B cells with phOx reactivity. Neither of the D-altered strains demonstrated the restriction in the VH/VL repertoire, the elimination of VH1 family-encoded antibodies, the focusing of the distribution of CDR-H3 lengths, or the selection for the normally dominant Ox1 clonotype, which all are hallmarks of the anti-phOx response in WT mice. These changes in clonal selection and expansion, as well as CSR indicate that the genetic constitution of the DH locus, which has been selected by evolution, can strongly influence the functional outcome of a TD humoral response.

No MeSH data available.


Related in: MedlinePlus

Variable gene usage of mAb of the thymus-dependent anti-phOx immune response in BALB/c wild-type and ΔD-DμFS and ΔD-iD DH mutant mice. Mice were immunized with the TD Ag phOx-CSA and mAb were prepared on day 7 (1°/7) and day 14 (1°/14°) after primary immunization and 3 days after secondary (2°), tertiary (3°), and quaternary (4°) immunizations, respectively. The characteristic attributes and utilized variable genes of these Ab are depicted in Tables S1 and S2 in Supplementary Material. Variability is calculated as the number of genes expressed by a group of antibodies divided by the number of monoclonal antibodies of this group and multiplied by 100. In BALB/c mice, IgM mAb (black bars) were only prepared early after primary immunization; see also legend to Figure 1. Data for IgG (white bars) are taken from two previous publications (9, 10). VH and VL variability is indicated for anti-phOx mAb of BALB/c wild-type mice (A,D), ΔD-DμFS mice (B,E), and ΔD-iD mice (C,F). (G–I) Expression of Ox1-idiotypic gene combination VH171/Vκ072 by monoclonal anti-phOx Ab from the TD response in BALB/c wild-type mice (G), ΔD-DμFS mice (H), and ΔD-iD mice (I). Data of BALB/c wild-type mice are taken from previous publications (5, 9, 10).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4128299&req=5

Figure 2: Variable gene usage of mAb of the thymus-dependent anti-phOx immune response in BALB/c wild-type and ΔD-DμFS and ΔD-iD DH mutant mice. Mice were immunized with the TD Ag phOx-CSA and mAb were prepared on day 7 (1°/7) and day 14 (1°/14°) after primary immunization and 3 days after secondary (2°), tertiary (3°), and quaternary (4°) immunizations, respectively. The characteristic attributes and utilized variable genes of these Ab are depicted in Tables S1 and S2 in Supplementary Material. Variability is calculated as the number of genes expressed by a group of antibodies divided by the number of monoclonal antibodies of this group and multiplied by 100. In BALB/c mice, IgM mAb (black bars) were only prepared early after primary immunization; see also legend to Figure 1. Data for IgG (white bars) are taken from two previous publications (9, 10). VH and VL variability is indicated for anti-phOx mAb of BALB/c wild-type mice (A,D), ΔD-DμFS mice (B,E), and ΔD-iD mice (C,F). (G–I) Expression of Ox1-idiotypic gene combination VH171/Vκ072 by monoclonal anti-phOx Ab from the TD response in BALB/c wild-type mice (G), ΔD-DμFS mice (H), and ΔD-iD mice (I). Data of BALB/c wild-type mice are taken from previous publications (5, 9, 10).

Mentions: In accordance with historical studies, in WT mice the anti-phOx response exhibited a drastic CSR-associated reduction in the variability of the VH and VL repertoires (10) (Figures 2A,D), especially in the class switched secondary response. No such reduction in VH or VL variability was observed in the D-altered ΔD-DμFS and ΔD-iD mice (Figures 2B,E and C,F). The classic secondary phOx response in WT mice is associated with a shift away from use of the VH1 family (10). However, in the D-altered mice hybridomas secreting VH1 family-encoded IgM and IgG mAb continued to be isolated at all phases of immune maturation (Tables S1 and S2 in Supplementary Material).


Clonal Progression during the T Cell-Dependent B Cell Antibody Response Depends on the Immunoglobulin DH Gene Segment Repertoire.

Trad A, Tanasa RI, Lange H, Zemlin M, Schroeder HW, Lemke H - Front Immunol (2014)

Variable gene usage of mAb of the thymus-dependent anti-phOx immune response in BALB/c wild-type and ΔD-DμFS and ΔD-iD DH mutant mice. Mice were immunized with the TD Ag phOx-CSA and mAb were prepared on day 7 (1°/7) and day 14 (1°/14°) after primary immunization and 3 days after secondary (2°), tertiary (3°), and quaternary (4°) immunizations, respectively. The characteristic attributes and utilized variable genes of these Ab are depicted in Tables S1 and S2 in Supplementary Material. Variability is calculated as the number of genes expressed by a group of antibodies divided by the number of monoclonal antibodies of this group and multiplied by 100. In BALB/c mice, IgM mAb (black bars) were only prepared early after primary immunization; see also legend to Figure 1. Data for IgG (white bars) are taken from two previous publications (9, 10). VH and VL variability is indicated for anti-phOx mAb of BALB/c wild-type mice (A,D), ΔD-DμFS mice (B,E), and ΔD-iD mice (C,F). (G–I) Expression of Ox1-idiotypic gene combination VH171/Vκ072 by monoclonal anti-phOx Ab from the TD response in BALB/c wild-type mice (G), ΔD-DμFS mice (H), and ΔD-iD mice (I). Data of BALB/c wild-type mice are taken from previous publications (5, 9, 10).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4128299&req=5

Figure 2: Variable gene usage of mAb of the thymus-dependent anti-phOx immune response in BALB/c wild-type and ΔD-DμFS and ΔD-iD DH mutant mice. Mice were immunized with the TD Ag phOx-CSA and mAb were prepared on day 7 (1°/7) and day 14 (1°/14°) after primary immunization and 3 days after secondary (2°), tertiary (3°), and quaternary (4°) immunizations, respectively. The characteristic attributes and utilized variable genes of these Ab are depicted in Tables S1 and S2 in Supplementary Material. Variability is calculated as the number of genes expressed by a group of antibodies divided by the number of monoclonal antibodies of this group and multiplied by 100. In BALB/c mice, IgM mAb (black bars) were only prepared early after primary immunization; see also legend to Figure 1. Data for IgG (white bars) are taken from two previous publications (9, 10). VH and VL variability is indicated for anti-phOx mAb of BALB/c wild-type mice (A,D), ΔD-DμFS mice (B,E), and ΔD-iD mice (C,F). (G–I) Expression of Ox1-idiotypic gene combination VH171/Vκ072 by monoclonal anti-phOx Ab from the TD response in BALB/c wild-type mice (G), ΔD-DμFS mice (H), and ΔD-iD mice (I). Data of BALB/c wild-type mice are taken from previous publications (5, 9, 10).
Mentions: In accordance with historical studies, in WT mice the anti-phOx response exhibited a drastic CSR-associated reduction in the variability of the VH and VL repertoires (10) (Figures 2A,D), especially in the class switched secondary response. No such reduction in VH or VL variability was observed in the D-altered ΔD-DμFS and ΔD-iD mice (Figures 2B,E and C,F). The classic secondary phOx response in WT mice is associated with a shift away from use of the VH1 family (10). However, in the D-altered mice hybridomas secreting VH1 family-encoded IgM and IgG mAb continued to be isolated at all phases of immune maturation (Tables S1 and S2 in Supplementary Material).

Bottom Line: We chose this antigen because studies of the humoral immune response to the hapten phOx were instrumental in the development of the current theoretical framework on which our understanding of the forces driving TD responses is based.Compared to WT, TD-induced humoral IgM as well as IgG antibody production in the D-altered ΔD-DμFS and ΔD-iD strains were significantly reduced.These changes in clonal selection and expansion, as well as CSR indicate that the genetic constitution of the DH locus, which has been selected by evolution, can strongly influence the functional outcome of a TD humoral response.

View Article: PubMed Central - PubMed

Affiliation: Biochemical Institute of the Medical Faculty of the Christian-Albrechts-University , Kiel , Germany.

ABSTRACT
The diversity of the third complementarity determining region of the IgH chain is constrained by natural selection of immunoglobulin diversity (DH) sequence. To test the functional significance of this constraint in the context of thymus-dependent (TD) immune responses, we immunized BALB/c mice with WT or altered DH sequence with 2-phenyloxazolone-coupled chicken serum albumin (phOx-CSA). We chose this antigen because studies of the humoral immune response to the hapten phOx were instrumental in the development of the current theoretical framework on which our understanding of the forces driving TD responses is based. To allow direct comparison, we used the classic approach of generating monoclonal Ab (mAb) from various stages of the immune response to phOx to assess the effect of changing the sequence of the DH on clonal expansion, class switching, and affinity maturation, which are hallmarks of TD responses. Compared to WT, TD-induced humoral IgM as well as IgG antibody production in the D-altered ΔD-DμFS and ΔD-iD strains were significantly reduced. An increased prevalence of IgM-producing hybridomas from late primary, secondary, and tertiary memory responses suggested either impaired class switch recombination (CSR) or impaired clonal expansion of class switched B cells with phOx reactivity. Neither of the D-altered strains demonstrated the restriction in the VH/VL repertoire, the elimination of VH1 family-encoded antibodies, the focusing of the distribution of CDR-H3 lengths, or the selection for the normally dominant Ox1 clonotype, which all are hallmarks of the anti-phOx response in WT mice. These changes in clonal selection and expansion, as well as CSR indicate that the genetic constitution of the DH locus, which has been selected by evolution, can strongly influence the functional outcome of a TD humoral response.

No MeSH data available.


Related in: MedlinePlus