# The 3D-Structures of the Immunoglobulin Heavy Chain Locus

> **NIH NIH R01** · UNIVERSITY OF CALIFORNIA, SAN DIEGO · 2024 · $380,263

## Abstract

Antibodies are generated by somatic recombination involving variable (V), diversity (D) and
joining (J) gene segments. During the previous grant cycle, we initiated studies aimed to
visualize VH-DHJH interactions in live cells. Using single color labeling we were able to track the
trajectories adopted by VH and DHJH elements. We found that VH and DHJH elements were
subjected to fractional Langevin motion, in which VH and DHJH elements bounce back and
forth in a spring-like fashion. However, since only a single genomic region was marked, this
approach did not permit us to directly monitor VH-DHJH interactions. To directly visualize VH-
DHJH encounters we developed a novel approach. We generated BCR-ABL transformed pro-
B cell lines that carried tandem arrays of wild-type TET-operator and mutant TET-operator
binding sites in the VH and DHJH regions, respectively. To mark the VH and DHJH regions
these cells were transduced with virus expressing WT-TET GFP and MUT-TET SNAP-TAG.
The results were surprising. VH-DHJH motion was severely sub-diffusive. We found that VH
regions were trapped in distinct chromatin configurations that were remarkably stable (<60
minutes). Only VH regions located nearby DHJH regions had a chance for a VH-DHJH
encounter. Comparison of simulated and experimental data suggested that such severely
sub-diffusive motion was imposed by geometric confinement (loop domains) and phase
separation/gelation (reversible cross-links within VH-DHJH and DH-JH loop domains). A caveat
of these studies is that they were performed using BCR-ABL transformed pro-B cells. Here we
propose to perform and extend these studies using primary pre-pro-B and pro-B cells.
Specifically, we would track VH-DHJH motion but now in primary B cell progenitors. We would
describe VH-DHJH motion in physical terms, including diffusion coefficients, scaling exponents,
velocities and spatial confinement. We would image across long-time scales to examine how
long VH and DHJH regions are trapped in distinct configurations and determine when (timing)
and how (speed) such configurations change. We would measure first-encounter times as well
as pairing times. We would examine whether transcriptional regulators and chromatin
remodelers modulate VH-DHJH motion and first-encounter times. We would identify critical
residues in RAG1 that would dictate pairing times. We would visualize VDJ recombination in
live cells. Data obtained from these experiments would reveal whether and how space and time
intersect to modulate the motion of paired coding and regulatory elements.

## Key facts

- **NIH application ID:** 10772153
- **Project number:** 5R01AI082850-15
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN DIEGO
- **Principal Investigator:** CORNELIS MURRE
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $380,263
- **Award type:** 5
- **Project period:** 2009-03-01 → 2026-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10772153

## Citation

> US National Institutes of Health, RePORTER application 10772153, The 3D-Structures of the Immunoglobulin Heavy Chain Locus (5R01AI082850-15). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10772153. Licensed CC0.

---

*[NIH grants dataset](/datasets/nih-grants) · CC0 1.0*