V(D)J Recombination and the Role of CTCF

Richard Wolfe; Xinhao Liu

T cell

T cell

1.A sort of lymphocytes

2.Related to Cell mediated immunity

3.Have T Cell recepor(Tcr) on the surface

B cell

B cell

1.A sort of lymphocytes

2.Related to humoral immunity

3.Have Bcr on the surface

4.Make antiboty

Light chain

Heavy chain

What is VDJ Recombination?

• V(D)J recombination is Somatic rearrangement of variable (V), diversity (D) and joining (J) regions of the genes that encode antigen receptors, leading to repertoire diversity of immunoglobulins and T cell receptors.

The structure of antigen receptor genes

Immunoglobulin

Heavy chain

Light chain

T cell receptor

Beta-chain

Alpha and d chain

Initiation of Recombination

• RAG(RAG recombinase) is a protein complex encoded by genes RAG1 and RAG2

HMGB1 or HMGB2 bind Spacers

Order during VDJ recombination

1. lgh and Tcrb are assembled with D to J recombination preceding V to DJ recombination

2. Immunoglobulin light chain(lgk or lgl) or Tcra is initiated.

Allelic exclusion: lg loci recombine full only in B cell lineage and TRC loci recombine only in T cells.

Question: How are the order, fidelity and allelic exclusion regulated during VDJ recombination?

Answer

Antigen receptor locus architecture and nuclear location work together with locus accessibility to ensure order, fidelity and allelic exclusion during the VDJ recombination

Character of Accessibility

V(D)J recombination correlates with numerous markers of open chromatin,germline transcription, activating histone modifications, nuclease accessibility and DNA hypomethylation, which indicate a tight connection between chromatin structure and targeting of recombination.

Character of Inaccessibility

1. A high nucleosome density, which affects direct positioning of RSSs on nucleosomes, cause steric constraints by nucleosome packing.

2. Deletion of enhancer accompanied by a loss of germline transcription

3. Deletion of promoter which has a more local effect.

4. antisense transcription

There are many other regulation of VDJ recomnination.

• CTCF, H3K9, H3K4, transcription factor, chromatin remodeling complex

CTCF as Repressor or Activator

• CTCF – CCCTC Binding Factor• Connects and separates different

chromatin domains through loop formation

• Mouse embryonic stem cells

• Four Types of Domains:

I. Formation of active domain

II. Formation of repressive domain

III. Formation of enhancer-promoter

IV. Formation of domain barrier between active and inactive chromatin regions

CTCF and the architecture of the Igh locus• Igh locus – V(D)J recombination• Heavy chain locus• Cohesin is a CTCF cofactor• Pre-proB stage is D-J recombination• ProB stage V-DJ recombination

a. CTCF and Cohesin binding sites• Cohesin binding at CTCF-sites more

specific pattern in proB stage

b. PAIR – Pax5-activated intergenic repeat elements

• PAIR elements bind with CTCF and mediate changes in locus conformation essential for V-DJ recombination

c. Pre-proB stage, CTCF bound to CB1/2 acts as an insulator, which stops antisense transcription and active chromatin from D spreading to V, thus limiting D-J recombination. In proB stage the barrier is released allowing V-DJ recombination

d. Model of CTCF mediated loops at preB stage assisting D-J recombination and at proB assisting V-DJ recombination

CTCF/Cohesin and the Igk and Tcra loci • Igk locus – VJ

recombination• Light chain locus• Tcra loci – T cell

a. CTCF binding more specific in preB cells

• Enlarged area shows CTCF site in Sis silencer as in igh locus

b. Approx. 60 CTCF binding sites of High and Low occupancy in V region

• Model shows CTCF loop separating enhancers from the V region

c. Again, CTCF binding site at TEA act to insulate the C-Jx region from the Dad1 area and Tcrd area

Conclusion

• CTCF and its cofactor, cohesin, have a fundamental role in regulating locus conformation and V(D)J rearrangement of the antigen receptor loci.

References

• Images:• Chaumeil J, Skok J: The role of CTCF in regulating

V(D)J recombination. Current Opinion in Immunology 2012, 24:153-159.

• Image of T cell, B cell and anti body are from google.• Image of recombination process are from David G.

Schatz* and Yanhong Ji, Recombination centres and the orchestration of V(D)J recombination. NATURE REVIEWS | IMMUNOLOGY 2011,11:251-263