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Silencing and nuclear repositioning of the lambda5 gene locus at the pre-B cell stage requires Aiolos and OBF-1.

Karnowski A, Cao C, Matthias G, Carotta S, Corcoran LM, Martensson IL, Skok JA, Matthias P - PLoS ONE (2008)

Bottom Line: However, when both factors are eliminated simultaneously a block is uncovered at the transition between pre-B and immature B cells, indicating that these proteins exert a critical function in developing B lymphocytes.Strikingly, developmentally regulated nuclear repositioning of the lambda5 gene is impaired in pre-B cells lacking OBF-1 and Aiolos.These studies uncover a novel role for OBF-1 and Aiolos in controlling the transcription and nuclear organization of genes involved in pre-BCR function.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute for Biomedical Research, Novartis Research Foundation, Basel, Switzerland.

ABSTRACT
The chromatin regulator Aiolos and the transcriptional coactivator OBF-1 have been implicated in regulating aspects of B cell maturation and activation. Mice lacking either of these factors have a largely normal early B cell development. However, when both factors are eliminated simultaneously a block is uncovered at the transition between pre-B and immature B cells, indicating that these proteins exert a critical function in developing B lymphocytes. In mice deficient for Aiolos and OBF-1, the numbers of immature B cells are reduced, small pre-BII cells are increased and a significant impairment in immunoglobulin light chain DNA rearrangement is observed. We identified genes whose expression is deregulated in the pre-B cell compartment of these mice. In particular, we found that components of the pre-BCR, such as the surrogate light chain genes lambda5 and VpreB, fail to be efficiently silenced in double-mutant mice. Strikingly, developmentally regulated nuclear repositioning of the lambda5 gene is impaired in pre-B cells lacking OBF-1 and Aiolos. These studies uncover a novel role for OBF-1 and Aiolos in controlling the transcription and nuclear organization of genes involved in pre-BCR function.

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Developmentally regulated nuclear repositioning of the λ5 locus is impaired in B lymphocytes lacking Aiolos or OBF-1.(A) Representative pictures of the DNA FISH analysis. Confocal sections of nuclei after DNA FISH are shown, combining the λ5 probe (red staining) with a probe detecting γ-satellite DNA (blue staining). The pictures show association of zero, one or both λ5 alleles with γ-satellite DNA. (B) Localization of the λ5 locus determined by DNA FISH analysis of bone marrow pre-BI (B220+ c-kit+ IgM−), small pre-BII (B220+ CD25+ IgM−), and splenic resting mature B cells (CD4− CD43− Ter− 119−). Percentages of nuclei with one (red bar), both (yellow bar) or neither (blue bar) λ5 allele associated with γ-satellite DNA in the indicated populations of wild type, single-and double-deficient mice. (C) Percentages of alleles located at the nuclear periphery in the indicated cell populations.
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pone-0003568-g006: Developmentally regulated nuclear repositioning of the λ5 locus is impaired in B lymphocytes lacking Aiolos or OBF-1.(A) Representative pictures of the DNA FISH analysis. Confocal sections of nuclei after DNA FISH are shown, combining the λ5 probe (red staining) with a probe detecting γ-satellite DNA (blue staining). The pictures show association of zero, one or both λ5 alleles with γ-satellite DNA. (B) Localization of the λ5 locus determined by DNA FISH analysis of bone marrow pre-BI (B220+ c-kit+ IgM−), small pre-BII (B220+ CD25+ IgM−), and splenic resting mature B cells (CD4− CD43− Ter− 119−). Percentages of nuclei with one (red bar), both (yellow bar) or neither (blue bar) λ5 allele associated with γ-satellite DNA in the indicated populations of wild type, single-and double-deficient mice. (C) Percentages of alleles located at the nuclear periphery in the indicated cell populations.

Mentions: In order to determine the impact of Aiolos and OBF-1 on this process, 3D DNA FISH was carried out on sorted pre-BI, small pre-BII as well as splenic mature resting B cells from wild type and mutant mice. A γ-satellite probe was used to detect centromeric clusters in conjunction with a VpreB/λ5 locus probe (Figure 6A). In wild-type pre-BI cells, the majority (65%) of cells has only one VpreB1/λ5 allele colocalizing with centromeric DNA and only a small proportion (13%) of the cells show both alleles positioned at the centromeric heterochromatin (Figure 6B). In the course of B cell development, a shift can be observed, as the VpreB1/λ5 alleles become progressively recruited to the centromeric DNA at the small pre-BII cell stage: at this stage a much greater proportion (48%) of the cells have both alleles associated with centromeric DNA and a minority of the cells (38%) have only one allele centromeric. Finally, in the majority of mature splenic B cells the VpreB1/λ5 alleles are no longer centromerically-associated. In addition, the recruitment to centromeric DNA is accompanied by translocation of the alleles to the nuclear periphery: while ca. 60% of VpreB/λ5 alleles are peripheral in wild type pre-BI cells, in small pre-BII or mature splenic B cells about 90% of the alleles are at the nuclear periphery (Figure 6C).


Silencing and nuclear repositioning of the lambda5 gene locus at the pre-B cell stage requires Aiolos and OBF-1.

Karnowski A, Cao C, Matthias G, Carotta S, Corcoran LM, Martensson IL, Skok JA, Matthias P - PLoS ONE (2008)

Developmentally regulated nuclear repositioning of the λ5 locus is impaired in B lymphocytes lacking Aiolos or OBF-1.(A) Representative pictures of the DNA FISH analysis. Confocal sections of nuclei after DNA FISH are shown, combining the λ5 probe (red staining) with a probe detecting γ-satellite DNA (blue staining). The pictures show association of zero, one or both λ5 alleles with γ-satellite DNA. (B) Localization of the λ5 locus determined by DNA FISH analysis of bone marrow pre-BI (B220+ c-kit+ IgM−), small pre-BII (B220+ CD25+ IgM−), and splenic resting mature B cells (CD4− CD43− Ter− 119−). Percentages of nuclei with one (red bar), both (yellow bar) or neither (blue bar) λ5 allele associated with γ-satellite DNA in the indicated populations of wild type, single-and double-deficient mice. (C) Percentages of alleles located at the nuclear periphery in the indicated cell populations.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2571989&req=5

pone-0003568-g006: Developmentally regulated nuclear repositioning of the λ5 locus is impaired in B lymphocytes lacking Aiolos or OBF-1.(A) Representative pictures of the DNA FISH analysis. Confocal sections of nuclei after DNA FISH are shown, combining the λ5 probe (red staining) with a probe detecting γ-satellite DNA (blue staining). The pictures show association of zero, one or both λ5 alleles with γ-satellite DNA. (B) Localization of the λ5 locus determined by DNA FISH analysis of bone marrow pre-BI (B220+ c-kit+ IgM−), small pre-BII (B220+ CD25+ IgM−), and splenic resting mature B cells (CD4− CD43− Ter− 119−). Percentages of nuclei with one (red bar), both (yellow bar) or neither (blue bar) λ5 allele associated with γ-satellite DNA in the indicated populations of wild type, single-and double-deficient mice. (C) Percentages of alleles located at the nuclear periphery in the indicated cell populations.
Mentions: In order to determine the impact of Aiolos and OBF-1 on this process, 3D DNA FISH was carried out on sorted pre-BI, small pre-BII as well as splenic mature resting B cells from wild type and mutant mice. A γ-satellite probe was used to detect centromeric clusters in conjunction with a VpreB/λ5 locus probe (Figure 6A). In wild-type pre-BI cells, the majority (65%) of cells has only one VpreB1/λ5 allele colocalizing with centromeric DNA and only a small proportion (13%) of the cells show both alleles positioned at the centromeric heterochromatin (Figure 6B). In the course of B cell development, a shift can be observed, as the VpreB1/λ5 alleles become progressively recruited to the centromeric DNA at the small pre-BII cell stage: at this stage a much greater proportion (48%) of the cells have both alleles associated with centromeric DNA and a minority of the cells (38%) have only one allele centromeric. Finally, in the majority of mature splenic B cells the VpreB1/λ5 alleles are no longer centromerically-associated. In addition, the recruitment to centromeric DNA is accompanied by translocation of the alleles to the nuclear periphery: while ca. 60% of VpreB/λ5 alleles are peripheral in wild type pre-BI cells, in small pre-BII or mature splenic B cells about 90% of the alleles are at the nuclear periphery (Figure 6C).

Bottom Line: However, when both factors are eliminated simultaneously a block is uncovered at the transition between pre-B and immature B cells, indicating that these proteins exert a critical function in developing B lymphocytes.Strikingly, developmentally regulated nuclear repositioning of the lambda5 gene is impaired in pre-B cells lacking OBF-1 and Aiolos.These studies uncover a novel role for OBF-1 and Aiolos in controlling the transcription and nuclear organization of genes involved in pre-BCR function.

View Article: PubMed Central - PubMed

Affiliation: Friedrich Miescher Institute for Biomedical Research, Novartis Research Foundation, Basel, Switzerland.

ABSTRACT
The chromatin regulator Aiolos and the transcriptional coactivator OBF-1 have been implicated in regulating aspects of B cell maturation and activation. Mice lacking either of these factors have a largely normal early B cell development. However, when both factors are eliminated simultaneously a block is uncovered at the transition between pre-B and immature B cells, indicating that these proteins exert a critical function in developing B lymphocytes. In mice deficient for Aiolos and OBF-1, the numbers of immature B cells are reduced, small pre-BII cells are increased and a significant impairment in immunoglobulin light chain DNA rearrangement is observed. We identified genes whose expression is deregulated in the pre-B cell compartment of these mice. In particular, we found that components of the pre-BCR, such as the surrogate light chain genes lambda5 and VpreB, fail to be efficiently silenced in double-mutant mice. Strikingly, developmentally regulated nuclear repositioning of the lambda5 gene is impaired in pre-B cells lacking OBF-1 and Aiolos. These studies uncover a novel role for OBF-1 and Aiolos in controlling the transcription and nuclear organization of genes involved in pre-BCR function.

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