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CTCF couples long-range loop extrusion and diffusional mechanism to mediate a diverse Igk repertoire

ABSTRACT: Dynamic genome folding plays a crucial role in cellular functions, including V(D)J recombination at the immunoglobulin kappa (Igκ) locus for antibody generation. This process involves frequent recombination between Jκ and Vκ gene segments over a 3.2 Mb region in both deletional and inversional orientations. While loop extrusion and diffusion are thought to facilitate Igκ recombination, their cooperative mechanisms remain unclear. Our study demonstrates CTCF as a key regulator coupling loop extrusion and diffusion to enhance Vκ recombination in both orientations across large genomic distances. We show that CTCF's N-terminus promotes long-range loop extrusion important for distal Vκ usage by stabilizing cohesin against Wapl release. Furthermore, CTCF's loop barrier function is essential for inversional Vκ usage via enabling diffusion. In CTCF N-terminus mutants, defects in inversional Vκ joining were not corrected by Wapl depletion alone but improved with targeted dCas9 blockade mimicking the CTCF barrier. These findings reveal that CTCF regulates immune diversity via diverse mechanisms and underscore its versatility in gene regulation.

INSTRUMENT(S):

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Liangliang Sun

LAB HEAD: Liangliang Sun

PROVIDER: PXD058006 | Pride | 2025-11-20

REPOSITORIES: Pride

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			Action	DRS
	Zhang_WT_IgG_1.raw	Raw
	Zhang_WT_IgG_2.raw	Raw
	Zhang_WT_IgG_3.raw	Raw
	Zhang_WT_SMC3IP_1.raw	Raw
	Zhang_WT_SMC3IP_2.raw	Raw

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CTCF couples long-range loop extrusion and diffusion to mediate a diverse Igk repertoire

Project description:Dynamic genome folding is important for V(D)J recombination at the immunoglobulin kappa (Igk) locus, which recombines Jk and Vk gene segments across a 3.2 Mb region in both deletional and inversional orientations. Chromatin loop extrusion and diffusion are considered two key mechanisms underlying Igk folding, but how they coordinate remains unclear. Here we show that CTCF is a key regulator coupling loop extrusion and diffusion during Igk V-J rearrangement, promoting recombination in both orientations across long genomic distances. Mechanistically, the CTCF N-terminus promotes long-range loop extrusion that facilitates distal Vk usage by stabilizing cohesin against WAPL release, and also forms loop barriers enabling chromatin diffusion for inversional Vk joining. In CTCF N-terminal-deficient B cells, defects in inversional Vk joining are not restored by WAPL depletion but are instead largely rescued by a dCas9-blockade targeted to the Vk-Jk intergenic region, mimicking the CTCF barrier. Our findings thus highlight how CTCF coordinates distinct genome-folding mechanisms through its dual roles in cohesin stabilization and extrusion barrier formation to ensure the generation of a diverse Igk repertoire.

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