Locus 'decontraction' and centromeric recruitment contribute to allelic exclusion of the immunoglobulin heavy-chain gene

Abstract

Allelic exclusion of immunoglobulin genes ensures the expression of a single antibody molecule in B cells through mostly unknown mechanisms. Large-scale contraction of the immunoglobulin heavy-chain (Igh) locus facilitates rearrangements between Igh variable (VH) and diversity gene segments in pro–B cells. Here we show that these long-range interactions are mediated by 'looping' of individual Igh subdomains. The Igk locus also underwent contraction by looping in small pre–B and immature B cells, demonstrating that immunoglobulin loci are in a contracted state in rearranging cells. Successful Igh recombination induced the rapid reversal of locus contraction in response to pre–B cell receptor signaling, which physically separated the distal VH genes from the proximal Igh domain, thus preventing further rearrangements. In the absence of locus contraction, only the four most proximal VH genes escaped allelic exclusion in immature μ-transgenic B lymphocytes. Pre–B cell receptor signaling also led to rapid repositioning of one Igh allele to repressive centromeric domains in response to downregulation of interleukin 7 signaling. These data link both locus 'decontraction' and centromeric recruitment to the establishment of allelic exclusion at the Igh locus.

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Figure 1: Decontraction of the Igh locus in large pre–B cells.
Figure 2: Contraction and decontraction of the Igk locus.
Figure 3: Contraction by looping of the Igh and Igk loci.
Figure 4: Monoallelic centromeric recruitment of the Igh locus during B cell development.
Figure 5: IL-7 signaling prevents centromeric recruitment of the Igh locus in splenic B cells.
Figure 6: Absence of centromeric recruitment and locus contraction of endogenous Igh alleles in μ-transgenic B cells.
Figure 7: Proximal VH-DJH rearrangements in M54 transgenic B cells.
Figure 8: The most proximal VH genes escape allelic exclusion in M54 transgenic B lymphocytes.

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Acknowledgements

We thank F. Batista for providing MD4 mice, and A. Corcoran and N. Mitchison for critically reviewing the manuscript. Supported by a Wellcome Trust University Award, Boehringer Ingelheim and the Austrian GEN-AU initiative (financed by the Bundesministerium für Bildung, Wissenschaft und Kultur).

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Correspondence to Meinrad Busslinger or Jane A Skok.

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Supplementary information

Supplementary Fig. 1

FACS sorting of B lymphocytes at different developmental stages. (PDF 813 kb)

Supplementary Fig. 2

Additional pictures with looping configurations of Igh and Igk alleles. (PDF 1334 kb)

Supplementary Fig. 3

The distal Vκ region of the Igk locus is recruited to centromeric domains. (PDF 1407 kb)

Supplementary Fig. 4

FACS sorting of M54 transgenic IgMa CD19+ B cells. (PDF 666 kb)

Supplementary Table 1

Separation of Igh and Igk probe signals in bone marrow B cells at distinct developmental stages and in activated splenic B cells. (PDF 60 kb)

Supplementary Table 2

Centromeric recruitment of Igh alleles in wild-type, MD4 and M54 cells at different stages of B cell development. (PDF 51 kb)

Supplementary Table 3

Reevaluation of the single-cell PCR data of large pre-B (preB-II) cells shown in Table 2 of ten Boekel et al. (Immunity 7, 357-368). (PDF 32 kb)

Supplementary Table 4

Centromeric recruitment of Ig loci in IL-7-treated splenic B cells and Igh probe separation in bone marrow B cells of M54 mice. (PDF 57 kb)

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Roldán, E., Fuxa, M., Chong, W. et al. Locus 'decontraction' and centromeric recruitment contribute to allelic exclusion of the immunoglobulin heavy-chain gene. Nat Immunol 6, 31–41 (2005). https://doi.org/10.1038/ni1150

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