Enhancers of the PAIR4 regulatory module promote distal VH gene recombination at the Igh locus
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE214869 | GEO | 2023/05/01
REPOSITORIES: GEO
Similar Datasets
Project description:VH-DJH recombination of the immunoglobulin heavy-chain (Igh) locus is temporally and spatially controlled during early B-cell development, and yet no regulatory elements other than the VH gene promoters have been identified throughout the entire 2.5-Mb VH gene cluster. Here we discovered novel regulatory sequences that are interspersed in the distal VH gene region. These conserved repeat elements were characterized by the presence of Pax5-dependent active chromatin, the binding of Pax5, E2A, CTCF and Rad21 as well as by Pax5-dependent antisense transcription in pro-B cells. The Pax5-activated intergenic repeat (PAIR) elements were no longer bound by Pax5 in pre-B and B cells consistent with the loss of antisense transcription, whereas E2A and CTCF interacted with PAIR elements throughout early B-cell development. The pro-B-cell-specific and Pax5-dependent activity of the PAIR elements suggests that they are involved in the regulation of distal VH-DJH recombination at the Igh locus. Analysis of chromatin and TF binding in rag2-/- and wt pro-B, DP T and Mature B cells. Chip-Seq of CTCF and Rad21. The provided data is in mm8 coordinates.
2011-02-11 | E-GEOD-27214 | biostudies-arrayexpress
Project description:VH-DJH recombination of the immunoglobulin heavy-chain (Igh) locus is temporally and spatially controlled during early B-cell development, and yet no regulatory elements other than the VH gene promoters have been identified throughout the entire 2.5-Mb VH gene cluster. Here we discovered novel regulatory sequences that are interspersed in the distal VH gene region. These conserved repeat elements were characterized by the presence of Pax5-dependent active chromatin, the binding of Pax5, E2A, CTCF and Rad21 as well as by Pax5-dependent antisense transcription in pro-B cells. The Pax5-activated intergenic repeat (PAIR) elements were no longer bound by Pax5 in pre-B and B cells consistent with the loss of antisense transcription, whereas E2A and CTCF interacted with PAIR elements throughout early B-cell development. The pro-B-cell-specific and Pax5-dependent activity of the PAIR elements suggests that they are involved in the regulation of distal VH-DJH recombination at the Igh locus. Analysis of chromatin and TF binding in rag2-/- and pax5-/- rag2-/- pro-B cells. Chip-Chip with one experiment for each antibody, 12 samples.
2011-02-11 | E-GEOD-27074 | biostudies-arrayexpress
Project description:VH-DJH recombination of the immunoglobulin heavy-chain (Igh) locus is temporally and spatially controlled during early B-cell development, and yet no regulatory elements other than the VH gene promoters have been identified throughout the entire 2.5-Mb VH gene cluster. Here we discovered novel regulatory sequences that are interspersed in the distal VH gene region. These conserved repeat elements were characterized by the presence of Pax5-dependent active chromatin, the binding of Pax5, E2A, CTCF and Rad21 as well as by Pax5-dependent antisense transcription in pro-B cells. The Pax5-activated intergenic repeat (PAIR) elements were no longer bound by Pax5 in pre-B and B cells consistent with the loss of antisense transcription, whereas E2A and CTCF interacted with PAIR elements throughout early B-cell development. The pro-B-cell-specific and Pax5-dependent activity of the PAIR elements suggests that they are involved in the regulation of distal VH-DJH recombination at the Igh locus.
2011-02-11 | GSE27214 | GEO
Project description:VH-DJH recombination of the immunoglobulin heavy-chain (Igh) locus is temporally and spatially controlled during early B-cell development, and yet no regulatory elements other than the VH gene promoters have been identified throughout the entire 2.5-Mb VH gene cluster. Here we discovered novel regulatory sequences that are interspersed in the distal VH gene region. These conserved repeat elements were characterized by the presence of Pax5-dependent active chromatin, the binding of Pax5, E2A, CTCF and Rad21 as well as by Pax5-dependent antisense transcription in pro-B cells. The Pax5-activated intergenic repeat (PAIR) elements were no longer bound by Pax5 in pre-B and B cells consistent with the loss of antisense transcription, whereas E2A and CTCF interacted with PAIR elements throughout early B-cell development. The pro-B-cell-specific and Pax5-dependent activity of the PAIR elements suggests that they are involved in the regulation of distal VH-DJH recombination at the Igh locus.
2011-02-11 | GSE27074 | GEO
Project description: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.
2020-05-11 | GSE142779 | GEO
Project description: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.
2020-05-11 | GSE142778 | GEO
Project description: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.
2020-05-11 | GSE142777 | GEO
Project description: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.
2020-05-11 | GSE142780 | GEO
Project description:Immunoglobulin heavy chain variable region exons are assembled in progenitor-B cells, from VH, D, and JH gene segments located in separate clusters across the Igh locus. RAG endonuclease orchestrates V(D)J recombination from a JH-based recombination center (RC). Cohesin-mediated extrusion of upstream chromatin past RC-bound RAG presents Ds for joining to JHs to form a DJHRC, from which RAG scans upstream for VHs to assemble VHDJH exons in an ordered recombination process. Cohesin-mediated chromatin loop extrusion is impeded by CTCF-bound elements (CBEs), with extrusion between two CBEs, particularly if convergently-oriented, forming chromatin-contact loops genome-wide. Igh has provocative CBE organization, with two divergently-oriented CBEs (CBE1 and CBE2) in the IGCR1 element between the VH and D/JH domains, dozens of VH domain CBEs convergent to CBE1, and 10 3'Igh-CBEs convergent to CBE2. IGCR1 CBEs segregate D/JH and VH domains by impeding RAG-scanning. After DJH formation, down-regulation of WAPL, a cohesin unloader, neutralizes IGCR1 and VH-domain CBEs, allowing RAG to scan the VH domain. To elucidate potential mechanisms underlying the developmental transition from D-to-JH to VH-to-DJH recombination and potential roles of IGCR1-based CBEs and 3'Igh-CBEs in regulating RAG-scanning, we tested effects of deleting or inverting IGCR1 or 3'Igh-CBEs in mice and/or progenitor-B cell lines. These studies revealed that normal IGCR1 CBE orientation augments its RAG-scanning impediment activity and that the 3'Igh-CBEs reinforce RC activity as a dynamic loop extrusion impediment, thereby, increasing its ability to support V(D)J recombination. Finally, our findings implicate a mechanism by which developmental WAPL down-regulation contributes to ordered V(D)J recombination.
2023-05-30 | GSE230251 | GEO
Project description:Immunoglobulin heavy chain variable region exons are assembled in progenitor-B cells, from VH, D, and JH gene segments located in separate clusters across the Igh locus. RAG endonuclease orchestrates V(D)J recombination from a JH-based recombination center (RC). Cohesin-mediated extrusion of upstream chromatin past RC-bound RAG presents Ds for joining to JHs to form a DJHRC, from which RAG scans upstream for VHs to assemble VHDJH exons in an ordered recombination process. Cohesin-mediated chromatin loop extrusion is impeded by CTCF-bound elements (CBEs), with extrusion between two CBEs, particularly if convergently-oriented, forming chromatin-contact loops genome-wide. Igh has provocative CBE organization, with two divergently-oriented CBEs (CBE1 and CBE2) in the IGCR1 element between the VH and D/JH domains, dozens of VH domain CBEs convergent to CBE1, and 10 3'Igh-CBEs convergent to CBE2. IGCR1 CBEs segregate D/JH and VH domains by impeding RAG-scanning. After DJH formation, down-regulation of WAPL, a cohesin unloader, neutralizes IGCR1 and VH-domain CBEs, allowing RAG to scan the VH domain. To elucidate potential mechanisms underlying the developmental transition from D-to-JH to VH-to-DJH recombination and potential roles of IGCR1-based CBEs and 3'Igh-CBEs in regulating RAG-scanning, we tested effects of deleting or inverting IGCR1 or 3'Igh-CBEs in mice and/or progenitor-B cell lines. These studies revealed that normal IGCR1 CBE orientation augments its RAG-scanning impediment activity and that the 3'Igh-CBEs reinforce RC activity as a dynamic loop extrusion impediment, thereby, increasing its ability to support V(D)J recombination. Finally, our findings implicate a mechanism by which developmental WAPL down-regulation contributes to ordered V(D)J recombination.
2023-05-30 | GSE230249 | GEO