title: CCCTC-binding factor (CTCF) and cohesin influence the genomic architecture of the Igh locus and antisense transcription in pro-B cells.
abstract: Compaction and looping of the ~2.5-Mb Igh locus during V(D)J rearrangement is essential to allow all V(H) genes to be brought in proximity with D(H)-J(H) segments to create a diverse antibody repertoire, but the proteins directly responsible for this are unknown. Because CCCTC-binding factor (CTCF) has been demonstrated to be involved in long-range chromosomal interactions, we hypothesized that CTCF may promote the contraction of the Igh locus. ChIP sequencing was performed on pro-B cells, revealing colocalization of CTCF and Rad21 binding at ~60 sites throughout the V(H) region and 2 other sites within the Igh locus. These numerous CTCF/cohesin sites potentially form the bases of the multiloop rosette structures at the Igh locus that compact during Ig heavy chain rearrangement. To test whether CTCF was involved in locus compaction, we used 3D-FISH to measure compaction in pro-B cells transduced with CTCF shRNA retroviruses. Reduction of CTCF binding resulted in a decrease in Igh locus compaction. Long-range interactions within the Igh locus were measured with the chromosomal conformation capture assay, revealing direct interactions between CTCF sites 5' of DFL16 and the 3' regulatory region, and also the intronic enhancer (Eμ), creating a D(H)-J(H)-Eμ-C(H) domain. Knockdown of CTCF also resulted in the increase of antisense transcription throughout the D(H) region and parts of the V(H) locus, suggesting a widespread regulatory role for CTCF. Together, our findings demonstrate that CTCF plays an important role in the 3D structure of the Igh locus and in the regulation of antisense germline transcription and that it contributes to the compaction of the Igh locus.
authors: Department of Immunology and Microbial Science, The Scripps Research Institute, La Jolla, CA 92037, USA.
authors: Stephanie C
authors: SC
authors: Degner
authors: Jiyoti
authors: J
authors: Verma-Gaur
authors: Timothy P
authors: TP
authors: Wong
authors: Claudia
authors: C
authors: Bossen
authors: G Michael
authors: GM
authors: Iverson
authors: Ali
authors: A
authors: Torkamani
authors: Christian
authors: C
authors: Vettermann
authors: Yin C
authors: YC
authors: Lin
authors: Zhongliang
authors: Z
authors: Ju
authors: Danae
authors: D
authors: Schulz
authors: Caroline S
authors: CS
authors: Murre
authors: Barbara K
authors: BK
authors: Birshtein
authors: Nicholas J
authors: NJ
authors: Schork
authors: Mark S
authors: MS
authors: Schlissel
authors: Roy
authors: R
authors: Riblet
authors: Cornelis
authors: C
authors: Murre
authors: Ann J
authors: AJ
authors: Feeney
references: Cell. 2009 Jun 26;137(7):1194-211
references: 19563753
references: Nat Immunol. 2005 Jan;6(1):31-41
references: 15580273
references: Genes Dev. 2005 Feb 1;19(3):322-7
references: 15687256
references: Genes Dev. 2007 May 15;21(10):1179-89
references: 17504937
references: Mol Cell. 2007 Jan 12;25(1):43-56
references: 17218270
references: Mol Cell Biol. 2005 Feb;25(4):1511-25
references: 15684400
references: Science. 2002 Apr 5;296(5565):158-62
references: 11935030
references: Proc Natl Acad Sci U S A. 2008 Jun 17;105(24):8309-14
references: 18550811
references: Proc Natl Acad Sci U S A. 2005 Oct 4;102(40):14362-7
references: 16186486
references: Immunity. 2001 Aug;15(2):187-99
references: 11520455
references: Cell. 2008 Feb 8;132(3):422-33
references: 18237772
references: Curr Opin Cell Biol. 2011 Jun;23(3):318-24
references: 21169003
references: Nat Immunol. 2003 Feb;4(2):124-31
references: 12496962
references: Curr Opin Genet Dev. 2007 Oct;17(5):400-7
references: 17913488
references: J Exp Med. 2007 Dec 24;204(13):3247-56
references: 18056289
references: Annu Rev Genet. 2009;43:525-58
references: 19886810
references: Immunity. 2011 Feb 25;34(2):175-87
references: 21349430
references: J Biol Chem. 2010 Mar 26;285(13):9327-9338
references: 20100833
references: Proc Natl Acad Sci U S A. 2010 Dec 21;107(51):22207-12
references: 21123744
references: Immunity. 2004 Mar;20(3):349-60
references: 15030778
references: Mol Cell Biol. 2007 Aug;27(15):5523-33
references: 17526723
references: Genes Dev. 2004 Feb 15;18(4):411-22
references: 15004008
references: Immunity. 2009 Oct 16;31(4):551-64
references: 19818655
references: Proc Natl Acad Sci U S A. 2004 Jun 8;101(23):8620-4
references: 15169959
references: Nature. 2008 Feb 14;451(7180):796-801
references: 18235444
references: Genes Dev. 2006 Sep 1;20(17):2349-54
references: 16951251
references: J Biol Chem. 2007 Nov 30;282(48):35169-78
references: 17921139
references: Nat Immunol. 2008 Aug;9(8):927-36
references: 18568028
references: Nat Protoc. 2007;2(7):1722-33
references: 17641637
references: J Immunol. 2009 Jan 1;182(1):44-8
references: 19109133
references: Mol Cell. 2007 Sep 7;27(5):842-50
references: 17803947
references: Proc Natl Acad Sci U S A. 2006 Jul 11;103(28):10684-9
references: 16815976
references: Immunity. 2007 Nov;27(5):711-22
references: 17980632
references: Cell. 2008 Apr 18;133(2):265-79
references: 18423198
references: Blood. 2003 Aug 15;102(4):1421-7
references: 12714490