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Series GSE142777 Query DataSets for GSE142777
Status Public on May 11, 2020
Title CTCF orchestrates long-range cohesin-driven V(D)J recombinational scanning [LAM-HTGTS]
Organism Mus musculus
Experiment type Other
Summary RAG endonuclease initiates V(D)J recombination in progenitor (pro)-B cells. Upon binding a recombination center (RC)-based JH, RAG scans upstream chromatin via loop extrusion, potentially mediated by cohesin, to locate Ds and assemble a DJH-based RC. CTCF looping factor-bound elements (CBEs) within the IGCR1 element upstream of the Ds impede RAG-scanning; but their inactivation allows scanning to proximal VHs where additional CBEs activate rearrangement and impede scanning any further upstream. Distal VH utilization is thought to involve diffusional RC access following large-scale Igh locus contraction. Here, we test the potential of linear RAG-scanning to mediate distal VH usage in G1-arrested, v-Abl-pro-B cell lines, which undergo robust D-to-JH rearrangement, but little VH-to-DJH rearrangement, presumably due to lack of locus contraction. Through an auxin-induced approach, we degrade cohesin-component Rad21 or CTCF in these G1-arrested lines, which maintain substantial viability throughout four-day experiments. Rad21 degradation eliminated all V(D)J recombination and RAG-scanning-associated interactions, except RC-located DQ52-to-JH joining in which synapsis occurs by diffusion11. Remarkably, while CTCF degradation suppressed most CBE-based chromatin interactions, it promoted robust RC interactions with, and VH-to-DJH joining of, distal VHs, with patterns similar to those of "locus-contracted" primary pro-B cells. Thus, down-modulation of CTCF-bound scanning-impediment activity promotes cohesin-driven RAG-scanning across the 2.7Mb Igh locus.
Overall design We performed LAM-HTGTS V(D)J-Seq, 3C-HTGTS, GRO-Seq and ChIP-Seq in v- Abl transformed pro-B cells and its various mutant derivatives to study roles of chromatin loop extrusion mediated RAG scanning in IgH V(D)J recombination.
Contributor(s) Alt FW, Zhaoqing B, Jiangman L
Citation(s) 32717742
Submission date Dec 31, 2019
Last update date Oct 08, 2020
Contact name Frederick W Alt
Organization name Boston Children's Hospital
Department PCMM
Lab Alt
Street address 1 Blackfan Circle
City Boston
State/province MA
ZIP/Postal code 02115
Country USA
Platforms (1)
GPL16417 Illumina MiSeq (Mus musculus)
Samples (110)
GSM4240097 JH4_Rad21degron_NT_a_rep1
GSM4240098 JH4_Rad21degron_NT_b_rep1
GSM4240099 JH4_Rad21degron_IAA_a_rep1
This SubSeries is part of SuperSeries:
GSE142781 CTCF orchestrates long-range cohesin-driven V(D)J recombinational scanning
BioProject PRJNA598326
SRA SRP239166

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SOFT formatted family file(s) SOFTHelp
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Series Matrix File(s) TXTHelp

Supplementary file Size Download File type/resource
GSE142777_RAW.tar 175.0 Mb (http)(custom) TAR (of TXT)
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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