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Bcl-6 mediates the germinal center B cell phenotype and lymphomagenesis through transcriptional repression of the DNA-damage sensor ATR

Nature Immunology volume 8, pages 705714 (2007) | Download Citation

Abstract

Antibody specificity and diversity is generated in B cells during germinal center maturation through clonal expansion while they undergo class-switch recombination and somatic hypermutation. Here we demonstrate that the transcriptional repressor Bcl-6 mediates this phenotype by directly repressing ATR in centroblasts and lymphoma cells. ATR is critical in replication and DNA damage–sensing checkpoints. Bcl-6 allowed B cells to evade ATR-mediated checkpoints and attenuated the response of the B cells to exogenous DNA damage. Repression of ATR was necessary and sufficient for those Bcl-6 activities. CD40 signaling 'rescued' B cells from those effects by disrupting the Bcl-6 transcription-repression complex on the promoter of the gene encoding ATR. Our data demonstrate a transcriptional regulatory loop whereby Bcl-6 mediates the centroblast phenotype through transient silencing of ATR.

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Acknowledgements

We thank S. Schreiber and K. Cimprich for FLAG-ATR constructs, and A. Follenzi for assistance in establishing high-efficiency lentiviral transduction of primary B cells. Supported by the Cancer Research Institute (S.M.R.), the National Cancer Center (J.M.P.), the National Cancer Institute (R01 CA104348 to A.M. and R01 CA100885 to M.C.), the Leukemia and Lymphoma Society (A.M. and M.C.), the Chemotherapy Foundation (A.M.), the Samuel Waxman Cancer Research Foundation (A.M.) and the G&P Foundation (A.M.).

Author information

Affiliations

  1. Department of Developmental and Molecular Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.

    • Stella Maris Ranuncolo
    • , Jose M Polo
    •  & Ari Melnick
  2. Hematology-Oncology Division, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.

    • Jamil Dierov
    •  & Martin Carroll
  3. NimbleGen Systems, Madison, Wisconsin 53711, USA.

    • Michael Singer
    •  & Roland Green
  4. Department of Molecular and Cell Biology, Division of Immunology, University of California, Berkeley, California 94720, USA.

    • Tracy Kuo
  5. Department of Molecular Genetics, Albert Einstein College of Medicine, Bronx, New York 10461, USA.

    • John Greally

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Contributions

S.M.R. designed and did experiments and wrote the paper; J.M.P. did chromatin immunoprecipitation (quantitative and on a chip); J.D. did 'comet' assays; M.S. did microarray experiments; T.K. developed critical reagents; J.G. designed microarray experiments; R.G. designed experiments; and M.C. and A.M. designed and conceived experiments and wrote the paper.

Competing interests

M.S. and R.G. are employees of NimbleGen Systems.

Corresponding authors

Correspondence to Martin Carroll or Ari Melnick.

Supplementary information

PDF files

  1. 1.

    Supplementary Fig. 1

    ChIP-on-chip of BCL6 on the CCL3 and GAPDH loci.

  2. 2.

    Supplementary Fig. 2

    Phenotypic characterization of primary B cell populations.

  3. 3.

    Supplementary Fig. 3

    BCL6 shRNA knockdown in centroblasts and DLBCL cells.

  4. 4.

    Supplementary Fig. 4

    Electroporation of a second BCL6 shRNA (shBCL6 II) mediates similar effects as shBCL6 I in centroblasts.

  5. 5.

    Supplementary Fig. 5

    BPI mediates similar effects as BCL6 shRNA in centroblasts.

  6. 6.

    Supplementary Fig. 6

    BCL6 shRNA II rescues ATR from BCL6-mediated repression and the BCL6 damage-sensing attenuation phenotype.

  7. 7.

    Supplementary Fig. 7

    A BCL6 transcriptional circuit mediates the germinal center B cell phenotype.

  8. 8.

    Supplementary Table 2

    shRNA and siRNA sequences used to deplete BCL6 and ATR (shRNA loop is in bold letters).

  9. 9.

    Supplementary Table 3

    Primers used for qPCR.

  10. 10.

    Supplementary Table 4

    Complete list of antibodies.

Excel files

  1. 1.

    Supplementary Table 1

    The table lists the coordinates of the gene loci, probe sequences, starting position of each probe, and log-ratio enrichment of DNA probes used for ChIP on chip on the ATR, CCL3 and GAPDH loci - including all three replicates as indicated (see accompanying Excel file).

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DOI

https://doi.org/10.1038/ni1478

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