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| Status |
Public on Nov 08, 2020 |
| Title |
Loop Extrusion Mediates Physiological Locus Contraction for V(D)J Recombination (3C-HTGTS) |
| Organism |
Mus musculus |
| Experiment type |
Other
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| Summary |
Immunoglobulin heavy chain locus (Igh) VH, D, and JH gene segments are developmentally assembled into V(D)J exons. RAG endonuclease initiates V(D)J recombination by binding a JH-recombination signal sequence (RSS) within a chromatin-based recombination center (RC) and then, in an orientation-dependent process, scans upstream D-containing chromatin presented by cohesin-mediated loop extrusion for convergent D-RSSs to initiate DJH-RC formation. In primary pro-B cells, 100s of upstream VH-associated RSSs, embedded in convergent orientation to the DJH-RC-RSS, gain proximity to the DJH-RC for VH-to-DJH joining via a mechanistically-undefined VH-locus contraction process. Here, we report that a 2.4 mega-base VH locus inversion in primary pro-B cells nearly abrogates rearrangements of normally convergent VH-RSSs and cryptic RSSs, even though locus contraction per se is maintained. Moreover, this inversion activated rearrangement of both cryptic VH-locus RSSs normally in the opposite orientation and, unexpectedly, of normally-oriented cryptic RSSs within multiple, sequential upstream convergent-CBE domains. Primary pro-B cells had significantly reduced transcription of Wapl, a cohesin-unloading factor, versus levels in v-Abl pro-B lines that lack marked locus contraction or distal VH rearrangements. Correspondingly, Wapl depletion in v-Abl lines activated VH-locus contraction and orientation-specific RAG-scanning across the VH-locus. Our findings indicate that locus contraction and physiological VH-to-DJH joining both are regulated via circumvention of CBE scanning impediments.
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| Overall design |
We performed 3C-HTGTS in BM pro-B cells and v- Abl transformed pro-B cells and its various mutant derivatives to study orientation-dependent linear RAG scanning and locus contraction in long-range cohesin-driven V(D)J recombination.
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| Contributor(s) |
Alt FW, Dai H, Hu H |
| Citation missing |
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| Submission date |
Jun 05, 2020 |
| Last update date |
Nov 08, 2020 |
| Contact name |
Frederick W Alt |
| E-mail(s) |
jianqiao.hu@childrens.harvard.edu
|
| 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 |
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| Platforms (1) |
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| Samples (19)
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| This SubSeries is part of SuperSeries: |
| GSE151910 |
Loop Extrusion Mediates Physiological Locus Contraction for V(D)J Recombination |
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| Relations |
| BioProject |
PRJNA637631 |
| SRA |
SRP266166 |
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