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Silencing of B cell receptor signals in human naive B cells.

Feldhahn N, Schwering I, Lee S, Wartenberg M, Klein F, Wang H, Zhou G, Wang SM, Rowley JD, Hescheler J, Krönke M, Rajewsky K, Küppers R, Müschen M - J. Exp. Med. (2002)

Bottom Line: In a functional assay, we show that down-regulation of inhibitory IgSF receptors and increased responsiveness to BCR stimulation in memory as compared with naive B cells at least partly results from interleukin (IL)-4 receptor signaling.Thus, LIRB1 and IL-4 may represent components of two nonoverlapping gene expression programs in naive and memory B cells, respectively: in naive B cells, a large group of inhibitory IgSF receptors can elevate the BCR signaling threshold to prevent these cells from premature activation and clonal expansion before GC-dependent affinity maturation.In memory B cells, facilitated responsiveness upon reencounter of the immunizing antigen may result from amplification of BCR signals at virtually all levels of signal transduction.

View Article: PubMed Central - PubMed

Affiliation: Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, 50931 Köln, Germany.

ABSTRACT
To identify changes in the regulation of B cell receptor (BCR) signals during the development of human B cells, we generated genome-wide gene expression profiles using the serial analysis of gene expression (SAGE) technique for CD34(+) hematopoietic stem cells (HSCs), pre-B cells, naive, germinal center (GC), and memory B cells. Comparing these SAGE profiles, genes encoding positive regulators of BCR signaling were expressed at consistently lower levels in naive B cells than in all other B cell subsets. Conversely, a large group of inhibitory signaling molecules, mostly belonging to the immunoglobulin superfamily (IgSF), were specifically or predominantly expressed in naive B cells. The quantitative differences observed by SAGE were corroborated by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometry. In a functional assay, we show that down-regulation of inhibitory IgSF receptors and increased responsiveness to BCR stimulation in memory as compared with naive B cells at least partly results from interleukin (IL)-4 receptor signaling. Conversely, activation or impairment of the inhibitory IgSF receptor LIRB1 affected BCR-dependent Ca(2+) mobilization only in naive but not memory B cells. Thus, LIRB1 and IL-4 may represent components of two nonoverlapping gene expression programs in naive and memory B cells, respectively: in naive B cells, a large group of inhibitory IgSF receptors can elevate the BCR signaling threshold to prevent these cells from premature activation and clonal expansion before GC-dependent affinity maturation. In memory B cells, facilitated responsiveness upon reencounter of the immunizing antigen may result from amplification of BCR signals at virtually all levels of signal transduction.

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Regulation of inhibitory IgSF receptors in memory B cells by IL-4. Naive and memory B cells were purified from peripheral blood and cultured either in medium alone, or with IL-4. In another set of experiments, memory B cells were cultured in the presence of a neutralizing anti–IL-4Rα antibody, which was added after 8 h of preincubation with IL-4. The left and center panels show amplification products of semiquantitative RT-PCR for positive regulatory PTKs (BLK, BTK, SYK), the linker molecule BLNK, the negative regulatory PTK CSK, the inhibitory PTPs SHIP and SHP1, and the inhibitory IgSF receptors LIRB1, LIRB2, LIRB5, SIgLec5, SIgLec8, and CD66. In the right panel, the genomic loci of these genes are indicated.
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fig5: Regulation of inhibitory IgSF receptors in memory B cells by IL-4. Naive and memory B cells were purified from peripheral blood and cultured either in medium alone, or with IL-4. In another set of experiments, memory B cells were cultured in the presence of a neutralizing anti–IL-4Rα antibody, which was added after 8 h of preincubation with IL-4. The left and center panels show amplification products of semiquantitative RT-PCR for positive regulatory PTKs (BLK, BTK, SYK), the linker molecule BLNK, the negative regulatory PTK CSK, the inhibitory PTPs SHIP and SHP1, and the inhibitory IgSF receptors LIRB1, LIRB2, LIRB5, SIgLec5, SIgLec8, and CD66. In the right panel, the genomic loci of these genes are indicated.

Mentions: As treatment of naive B cells with IL-4 had no effect on the expression of genes related to BCR signaling (see above; see Fig. 5), modulation of BCR signals by IL-4 was studied in memory B cells only. Memory B cells from four healthy donors were purified and cultured in supernatant from LPS-stimulated PBMCs for 24 h in the presence or absence of human recombinant IL-4 or an inhibitory anti–IL-4Rα antibody (Genzyme). Changes of cytosolic Ca2+ concentrations upon BCR engagement were measured and analyzed as described above.


Silencing of B cell receptor signals in human naive B cells.

Feldhahn N, Schwering I, Lee S, Wartenberg M, Klein F, Wang H, Zhou G, Wang SM, Rowley JD, Hescheler J, Krönke M, Rajewsky K, Küppers R, Müschen M - J. Exp. Med. (2002)

Regulation of inhibitory IgSF receptors in memory B cells by IL-4. Naive and memory B cells were purified from peripheral blood and cultured either in medium alone, or with IL-4. In another set of experiments, memory B cells were cultured in the presence of a neutralizing anti–IL-4Rα antibody, which was added after 8 h of preincubation with IL-4. The left and center panels show amplification products of semiquantitative RT-PCR for positive regulatory PTKs (BLK, BTK, SYK), the linker molecule BLNK, the negative regulatory PTK CSK, the inhibitory PTPs SHIP and SHP1, and the inhibitory IgSF receptors LIRB1, LIRB2, LIRB5, SIgLec5, SIgLec8, and CD66. In the right panel, the genomic loci of these genes are indicated.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2193982&req=5

fig5: Regulation of inhibitory IgSF receptors in memory B cells by IL-4. Naive and memory B cells were purified from peripheral blood and cultured either in medium alone, or with IL-4. In another set of experiments, memory B cells were cultured in the presence of a neutralizing anti–IL-4Rα antibody, which was added after 8 h of preincubation with IL-4. The left and center panels show amplification products of semiquantitative RT-PCR for positive regulatory PTKs (BLK, BTK, SYK), the linker molecule BLNK, the negative regulatory PTK CSK, the inhibitory PTPs SHIP and SHP1, and the inhibitory IgSF receptors LIRB1, LIRB2, LIRB5, SIgLec5, SIgLec8, and CD66. In the right panel, the genomic loci of these genes are indicated.
Mentions: As treatment of naive B cells with IL-4 had no effect on the expression of genes related to BCR signaling (see above; see Fig. 5), modulation of BCR signals by IL-4 was studied in memory B cells only. Memory B cells from four healthy donors were purified and cultured in supernatant from LPS-stimulated PBMCs for 24 h in the presence or absence of human recombinant IL-4 or an inhibitory anti–IL-4Rα antibody (Genzyme). Changes of cytosolic Ca2+ concentrations upon BCR engagement were measured and analyzed as described above.

Bottom Line: In a functional assay, we show that down-regulation of inhibitory IgSF receptors and increased responsiveness to BCR stimulation in memory as compared with naive B cells at least partly results from interleukin (IL)-4 receptor signaling.Thus, LIRB1 and IL-4 may represent components of two nonoverlapping gene expression programs in naive and memory B cells, respectively: in naive B cells, a large group of inhibitory IgSF receptors can elevate the BCR signaling threshold to prevent these cells from premature activation and clonal expansion before GC-dependent affinity maturation.In memory B cells, facilitated responsiveness upon reencounter of the immunizing antigen may result from amplification of BCR signals at virtually all levels of signal transduction.

View Article: PubMed Central - PubMed

Affiliation: Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, 50931 Köln, Germany.

ABSTRACT
To identify changes in the regulation of B cell receptor (BCR) signals during the development of human B cells, we generated genome-wide gene expression profiles using the serial analysis of gene expression (SAGE) technique for CD34(+) hematopoietic stem cells (HSCs), pre-B cells, naive, germinal center (GC), and memory B cells. Comparing these SAGE profiles, genes encoding positive regulators of BCR signaling were expressed at consistently lower levels in naive B cells than in all other B cell subsets. Conversely, a large group of inhibitory signaling molecules, mostly belonging to the immunoglobulin superfamily (IgSF), were specifically or predominantly expressed in naive B cells. The quantitative differences observed by SAGE were corroborated by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometry. In a functional assay, we show that down-regulation of inhibitory IgSF receptors and increased responsiveness to BCR stimulation in memory as compared with naive B cells at least partly results from interleukin (IL)-4 receptor signaling. Conversely, activation or impairment of the inhibitory IgSF receptor LIRB1 affected BCR-dependent Ca(2+) mobilization only in naive but not memory B cells. Thus, LIRB1 and IL-4 may represent components of two nonoverlapping gene expression programs in naive and memory B cells, respectively: in naive B cells, a large group of inhibitory IgSF receptors can elevate the BCR signaling threshold to prevent these cells from premature activation and clonal expansion before GC-dependent affinity maturation. In memory B cells, facilitated responsiveness upon reencounter of the immunizing antigen may result from amplification of BCR signals at virtually all levels of signal transduction.

Show MeSH
Related in: MedlinePlus