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Expression levels of B cell surface immunoglobulin regulate efficiency of allelic exclusion and size of autoreactive B-1 cell compartment.

Watanabe N, Nisitani S, Ikuta K, Suzuki M, Chiba T, Honjo T - J. Exp. Med. (1999)

Bottom Line: Surface-expressed immunoglobulin (Ig) has been shown to have a critical role in allelic exclusion of Ig heavy (H) and light (L) chains.These results suggest that there is a threshold of the B cell receptor expression level that induces allelic exclusion.By contrast, homozygosity of the H chain alone Tg line, which does not recognize self-antigens, did not increase the size of the peritoneal B-1 subset.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Chemistry, Faculty of Medicine, Kyoto University, Yoshida, Sakyo-ku, Kyoto 606-8501, Japan.

ABSTRACT
Surface-expressed immunoglobulin (Ig) has been shown to have a critical role in allelic exclusion of Ig heavy (H) and light (L) chains. Although various degrees of suppression of endogenous Ig expression are observed in Ig transgenic (Tg) mice, it was not clear whether this difference is due to different onsets of Tg expression or to different levels of Tg expression, which are obviously affected by integration sites of the transgene. In this study we generated antierythrocyte antibody Tg mice that carry tandem joined H and L chain transgenes (H+L) and confirmed that homozygosity of the transgene loci enhances the level of transgene expression as compared with heterozygosity. Suppression of endogenous H and L chain gene expression was stronger in homozygous than in heterozygous Tg mice. Similar results were obtained in control Tg mice carrying the H chain only. These results suggest that there is a threshold of the B cell receptor expression level that induces allelic exclusion. In addition, despite the same B cell receptor specificity, the size of Tg autoreactive B-1 cell compartment in the peritoneal cavity is larger in homozygous than in heterozygous mice, although the number of the Tg B-2 cell subset decreased in the spleen and bone marrow of homozygous Tg mice as compared with heterozygous Tg mice. By contrast, homozygosity of the H chain alone Tg line, which does not recognize self-antigens, did not increase the size of the peritoneal B-1 subset. These results suggest that the size of the B-1 cell subset in the Tg mice may depend on strength of signals through B cell receptors triggered by self-antigens.

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Tg H chain expression in B cells of heterozygous or homozygous H+L and H3 Tg mice. Cells were isolated from bone marrow (A) and spleen (B) of indicated mice at 10–14 wk of age and stained with FITC-conjugated antiallotypic antibody for IgMa (Tg) and PE-conjugated antiallotypic antibody for IgMb (endogenous). Control stainings of IgMa (BALB/c) and IgMb (C57BL/6) spleen cells are shown. The percentages of cells of each gated region in total viable cells are indicated. Total numbers of the cells with endogenous H chains in heterozygous or homozygous H+L5, H+L6, and H3 Tg mice are shown (C). Spleen cells were stained and analyzed from at least three mice of each Tg line. Numbers of indicated cells were calculated by (percentage of indicated cells in viable cells) × (No. of viable cells). Results are expressed as mean ± SD.
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Figure 2: Tg H chain expression in B cells of heterozygous or homozygous H+L and H3 Tg mice. Cells were isolated from bone marrow (A) and spleen (B) of indicated mice at 10–14 wk of age and stained with FITC-conjugated antiallotypic antibody for IgMa (Tg) and PE-conjugated antiallotypic antibody for IgMb (endogenous). Control stainings of IgMa (BALB/c) and IgMb (C57BL/6) spleen cells are shown. The percentages of cells of each gated region in total viable cells are indicated. Total numbers of the cells with endogenous H chains in heterozygous or homozygous H+L5, H+L6, and H3 Tg mice are shown (C). Spleen cells were stained and analyzed from at least three mice of each Tg line. Numbers of indicated cells were calculated by (percentage of indicated cells in viable cells) × (No. of viable cells). Results are expressed as mean ± SD.

Mentions: To test whether inhibition of endogenous H chain expression is stronger in homozygous than in heterozygous mice, we analyzed lymphocytes in bone marrow and spleen of H+L5, H+L6, and H3 mice by flow cytometry. We stained bone marrow and spleen cells with antiallotypic mAbs (which can clearly distinguish IgM of the control mice; Fig. 2A and Fig. B) for IgMa (Tg) and IgMb (endogenous). In all heterozygous Tg mice, the numbers of IgMa+IgMb+ cells (allelic inclusion) were small but significant, whereas these cells almost completely disappeared in all homozygous Tg mice. This conclusion is further confirmed by the finding that both bone marrow and spleen B cells with only endogenous H chains (IgMa−IgMb+) decreased in homozygous Tg mice as compared with heterozygous Tg mice (Fig. 2A and Fig. B). In all lines of Tg mice, total numbers of endogenous H chain–expressing (IgMb+) spleen cells were lower in homozygous than in heterozygous mice (Fig. 2 C). These results indicate that inhibition of endogenous H chain expression (allelic exclusion) is stronger in homozygous than in heterozygous mice. Taken together, higher levels of Ig transgene expression appear to cause stronger inhibition of endogenous H chain expression, suggesting that a certain level of surface Ig expression is required for allelic exclusion of the H chain locus.


Expression levels of B cell surface immunoglobulin regulate efficiency of allelic exclusion and size of autoreactive B-1 cell compartment.

Watanabe N, Nisitani S, Ikuta K, Suzuki M, Chiba T, Honjo T - J. Exp. Med. (1999)

Tg H chain expression in B cells of heterozygous or homozygous H+L and H3 Tg mice. Cells were isolated from bone marrow (A) and spleen (B) of indicated mice at 10–14 wk of age and stained with FITC-conjugated antiallotypic antibody for IgMa (Tg) and PE-conjugated antiallotypic antibody for IgMb (endogenous). Control stainings of IgMa (BALB/c) and IgMb (C57BL/6) spleen cells are shown. The percentages of cells of each gated region in total viable cells are indicated. Total numbers of the cells with endogenous H chains in heterozygous or homozygous H+L5, H+L6, and H3 Tg mice are shown (C). Spleen cells were stained and analyzed from at least three mice of each Tg line. Numbers of indicated cells were calculated by (percentage of indicated cells in viable cells) × (No. of viable cells). Results are expressed as mean ± SD.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2195600&req=5

Figure 2: Tg H chain expression in B cells of heterozygous or homozygous H+L and H3 Tg mice. Cells were isolated from bone marrow (A) and spleen (B) of indicated mice at 10–14 wk of age and stained with FITC-conjugated antiallotypic antibody for IgMa (Tg) and PE-conjugated antiallotypic antibody for IgMb (endogenous). Control stainings of IgMa (BALB/c) and IgMb (C57BL/6) spleen cells are shown. The percentages of cells of each gated region in total viable cells are indicated. Total numbers of the cells with endogenous H chains in heterozygous or homozygous H+L5, H+L6, and H3 Tg mice are shown (C). Spleen cells were stained and analyzed from at least three mice of each Tg line. Numbers of indicated cells were calculated by (percentage of indicated cells in viable cells) × (No. of viable cells). Results are expressed as mean ± SD.
Mentions: To test whether inhibition of endogenous H chain expression is stronger in homozygous than in heterozygous mice, we analyzed lymphocytes in bone marrow and spleen of H+L5, H+L6, and H3 mice by flow cytometry. We stained bone marrow and spleen cells with antiallotypic mAbs (which can clearly distinguish IgM of the control mice; Fig. 2A and Fig. B) for IgMa (Tg) and IgMb (endogenous). In all heterozygous Tg mice, the numbers of IgMa+IgMb+ cells (allelic inclusion) were small but significant, whereas these cells almost completely disappeared in all homozygous Tg mice. This conclusion is further confirmed by the finding that both bone marrow and spleen B cells with only endogenous H chains (IgMa−IgMb+) decreased in homozygous Tg mice as compared with heterozygous Tg mice (Fig. 2A and Fig. B). In all lines of Tg mice, total numbers of endogenous H chain–expressing (IgMb+) spleen cells were lower in homozygous than in heterozygous mice (Fig. 2 C). These results indicate that inhibition of endogenous H chain expression (allelic exclusion) is stronger in homozygous than in heterozygous mice. Taken together, higher levels of Ig transgene expression appear to cause stronger inhibition of endogenous H chain expression, suggesting that a certain level of surface Ig expression is required for allelic exclusion of the H chain locus.

Bottom Line: Surface-expressed immunoglobulin (Ig) has been shown to have a critical role in allelic exclusion of Ig heavy (H) and light (L) chains.These results suggest that there is a threshold of the B cell receptor expression level that induces allelic exclusion.By contrast, homozygosity of the H chain alone Tg line, which does not recognize self-antigens, did not increase the size of the peritoneal B-1 subset.

View Article: PubMed Central - PubMed

Affiliation: Department of Medical Chemistry, Faculty of Medicine, Kyoto University, Yoshida, Sakyo-ku, Kyoto 606-8501, Japan.

ABSTRACT
Surface-expressed immunoglobulin (Ig) has been shown to have a critical role in allelic exclusion of Ig heavy (H) and light (L) chains. Although various degrees of suppression of endogenous Ig expression are observed in Ig transgenic (Tg) mice, it was not clear whether this difference is due to different onsets of Tg expression or to different levels of Tg expression, which are obviously affected by integration sites of the transgene. In this study we generated antierythrocyte antibody Tg mice that carry tandem joined H and L chain transgenes (H+L) and confirmed that homozygosity of the transgene loci enhances the level of transgene expression as compared with heterozygosity. Suppression of endogenous H and L chain gene expression was stronger in homozygous than in heterozygous Tg mice. Similar results were obtained in control Tg mice carrying the H chain only. These results suggest that there is a threshold of the B cell receptor expression level that induces allelic exclusion. In addition, despite the same B cell receptor specificity, the size of Tg autoreactive B-1 cell compartment in the peritoneal cavity is larger in homozygous than in heterozygous mice, although the number of the Tg B-2 cell subset decreased in the spleen and bone marrow of homozygous Tg mice as compared with heterozygous Tg mice. By contrast, homozygosity of the H chain alone Tg line, which does not recognize self-antigens, did not increase the size of the peritoneal B-1 subset. These results suggest that the size of the B-1 cell subset in the Tg mice may depend on strength of signals through B cell receptors triggered by self-antigens.

Show MeSH
Related in: MedlinePlus