Limits...
Mimicry of pre-B cell receptor signaling by activation of the tyrosine kinase Blk.

Tretter T, Ross AE, Dordai DI, Desiderio S - J. Exp. Med. (2003)

Bottom Line: Here the Src-related kinase Blk is shown to effect functions associated with the pre-BCR.These alterations were accompanied by tyrosine phosphorylation of immunoglobulin beta and Syk, as well as changes in gene expression consistent with developmental maturation.Thus, sustained activation of Blk induces responses normally associated with the pre-BCR.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

ABSTRACT
During B lymphoid ontogeny, assembly of the pre-B cell receptor (BCR) is a principal developmental checkpoint at which several Src-related kinases may play redundant roles. Here the Src-related kinase Blk is shown to effect functions associated with the pre-BCR. B lymphoid expression of an active Blk mutant caused proliferation of B progenitor cells and enhanced responsiveness of these cells to interleukin 7. In mice lacking a functional pre-BCR, active Blk supported maturation beyond the pro-B cell stage, suppressed VH to DJH rearrangement, relieved selection for productive heavy chain rearrangement, and stimulated kappa rearrangement. These alterations were accompanied by tyrosine phosphorylation of immunoglobulin beta and Syk, as well as changes in gene expression consistent with developmental maturation. Thus, sustained activation of Blk induces responses normally associated with the pre-BCR.

Show MeSH
Appearance of cells bearing a B progenitor phenotype in spleens of Blk(Y495F) transgenic mice. (A) Flow cytometric analysis. Single cell suspensions from spleens of transgenic (left) or nontransgenic (right) littermates (3–5 wk old) were stained with antibodies to the indicated markers. Plots of BP-1, CD22, or CD43 versus IgM (bottom three pairs) are gated on B220+ cells. Numbers indicate the percentage of cells in the corresponding quadrant. (B) Peripheral expression of immature B cell markers in Blk(Y495F) transgenic mice. RNA was prepared from spleens of nontransgenic (lanes 1–3) or transgenic (lanes 4–6) littermates and transcripts encoding RAG-2, terminal nucleotidyl transferase (TdT), VpreB, or actin were detected by RT-PCR. Products were diluted as indicated above, fractionated by gel electrophoresis, and detected by staining with ethidium bromide.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2194155&req=5

fig5: Appearance of cells bearing a B progenitor phenotype in spleens of Blk(Y495F) transgenic mice. (A) Flow cytometric analysis. Single cell suspensions from spleens of transgenic (left) or nontransgenic (right) littermates (3–5 wk old) were stained with antibodies to the indicated markers. Plots of BP-1, CD22, or CD43 versus IgM (bottom three pairs) are gated on B220+ cells. Numbers indicate the percentage of cells in the corresponding quadrant. (B) Peripheral expression of immature B cell markers in Blk(Y495F) transgenic mice. RNA was prepared from spleens of nontransgenic (lanes 1–3) or transgenic (lanes 4–6) littermates and transcripts encoding RAG-2, terminal nucleotidyl transferase (TdT), VpreB, or actin were detected by RT-PCR. Products were diluted as indicated above, fractionated by gel electrophoresis, and detected by staining with ethidium bromide.

Mentions: B cell progenitors can accumulate in secondary lymphoid tissues under conditions of stress or polyclonal activation (37, 38), as well as in RAG- or heavy chain–deficient mice expressing activated Ras in the B lymphoid compartment (39, 40). This also occurred when Blk(Y495F) was expressed in the B lineage. Although overall splenic structure and cellularity were not significantly affected by the transgene, a substantial decrease in the percentage of B220+ cells expressing IgM was seen in spleens of 3–5-wk-old transgenic animals relative to nontransgenic littermates (16.9 ± 2.4 in transgenic animals, 51.6 ± 1.7 in nontransgenic controls; Fig. 5 A and Table S2, available at http://www.jem.org/cgi/content/full/jem.20030729/DC1). Most remaining B lymphoid cells were B220lo, CD43int, CD22int, and BP-1high, suggesting that they were derived from the B progenitor population that accumulates in bone marrow. Consistent with this interpretation, splenic RNA from transgenic mice contained transcripts corresponding to RAG-2, TdT, VpreB (Fig. 5 B), and λ5 (not depicted), whose expression is characteristic of lymphoid progenitors. The B progenitors found in the spleens of young transgenic mice were polyclonal (unpublished data), in contrast to the clonal tumors that arise in these mice after 6–12 mo. These observations suggest that in Blk(Y495F) transgenic mice, B cell progenitors emigrate to peripheral lymphoid organs in the absence of BCR expression.


Mimicry of pre-B cell receptor signaling by activation of the tyrosine kinase Blk.

Tretter T, Ross AE, Dordai DI, Desiderio S - J. Exp. Med. (2003)

Appearance of cells bearing a B progenitor phenotype in spleens of Blk(Y495F) transgenic mice. (A) Flow cytometric analysis. Single cell suspensions from spleens of transgenic (left) or nontransgenic (right) littermates (3–5 wk old) were stained with antibodies to the indicated markers. Plots of BP-1, CD22, or CD43 versus IgM (bottom three pairs) are gated on B220+ cells. Numbers indicate the percentage of cells in the corresponding quadrant. (B) Peripheral expression of immature B cell markers in Blk(Y495F) transgenic mice. RNA was prepared from spleens of nontransgenic (lanes 1–3) or transgenic (lanes 4–6) littermates and transcripts encoding RAG-2, terminal nucleotidyl transferase (TdT), VpreB, or actin were detected by RT-PCR. Products were diluted as indicated above, fractionated by gel electrophoresis, and detected by staining with ethidium bromide.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2194155&req=5

fig5: Appearance of cells bearing a B progenitor phenotype in spleens of Blk(Y495F) transgenic mice. (A) Flow cytometric analysis. Single cell suspensions from spleens of transgenic (left) or nontransgenic (right) littermates (3–5 wk old) were stained with antibodies to the indicated markers. Plots of BP-1, CD22, or CD43 versus IgM (bottom three pairs) are gated on B220+ cells. Numbers indicate the percentage of cells in the corresponding quadrant. (B) Peripheral expression of immature B cell markers in Blk(Y495F) transgenic mice. RNA was prepared from spleens of nontransgenic (lanes 1–3) or transgenic (lanes 4–6) littermates and transcripts encoding RAG-2, terminal nucleotidyl transferase (TdT), VpreB, or actin were detected by RT-PCR. Products were diluted as indicated above, fractionated by gel electrophoresis, and detected by staining with ethidium bromide.
Mentions: B cell progenitors can accumulate in secondary lymphoid tissues under conditions of stress or polyclonal activation (37, 38), as well as in RAG- or heavy chain–deficient mice expressing activated Ras in the B lymphoid compartment (39, 40). This also occurred when Blk(Y495F) was expressed in the B lineage. Although overall splenic structure and cellularity were not significantly affected by the transgene, a substantial decrease in the percentage of B220+ cells expressing IgM was seen in spleens of 3–5-wk-old transgenic animals relative to nontransgenic littermates (16.9 ± 2.4 in transgenic animals, 51.6 ± 1.7 in nontransgenic controls; Fig. 5 A and Table S2, available at http://www.jem.org/cgi/content/full/jem.20030729/DC1). Most remaining B lymphoid cells were B220lo, CD43int, CD22int, and BP-1high, suggesting that they were derived from the B progenitor population that accumulates in bone marrow. Consistent with this interpretation, splenic RNA from transgenic mice contained transcripts corresponding to RAG-2, TdT, VpreB (Fig. 5 B), and λ5 (not depicted), whose expression is characteristic of lymphoid progenitors. The B progenitors found in the spleens of young transgenic mice were polyclonal (unpublished data), in contrast to the clonal tumors that arise in these mice after 6–12 mo. These observations suggest that in Blk(Y495F) transgenic mice, B cell progenitors emigrate to peripheral lymphoid organs in the absence of BCR expression.

Bottom Line: Here the Src-related kinase Blk is shown to effect functions associated with the pre-BCR.These alterations were accompanied by tyrosine phosphorylation of immunoglobulin beta and Syk, as well as changes in gene expression consistent with developmental maturation.Thus, sustained activation of Blk induces responses normally associated with the pre-BCR.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.

ABSTRACT
During B lymphoid ontogeny, assembly of the pre-B cell receptor (BCR) is a principal developmental checkpoint at which several Src-related kinases may play redundant roles. Here the Src-related kinase Blk is shown to effect functions associated with the pre-BCR. B lymphoid expression of an active Blk mutant caused proliferation of B progenitor cells and enhanced responsiveness of these cells to interleukin 7. In mice lacking a functional pre-BCR, active Blk supported maturation beyond the pro-B cell stage, suppressed VH to DJH rearrangement, relieved selection for productive heavy chain rearrangement, and stimulated kappa rearrangement. These alterations were accompanied by tyrosine phosphorylation of immunoglobulin beta and Syk, as well as changes in gene expression consistent with developmental maturation. Thus, sustained activation of Blk induces responses normally associated with the pre-BCR.

Show MeSH