CTCF-binding elements 1 and 2 in the Igh intergenic control region cooperatively regulate V(D)J recombination

Area of Science:

• Immunology
• Molecular Biology
• Genetics

Background:

• B-lymphocyte differentiation involves immunoglobulin heavy chain (IgH) variable region exon assembly from V, D, and J gene segments.
• The mouse IgH locus contains hundreds of V(H) genes, regulated by the intergenic control region 1 (IGCR1).
• IGCR1 controls IgH V(D)J recombination, influencing developmental order, lineage specificity, and B-cell receptor repertoire diversity.

Purpose of the Study:

• To investigate the individual and cooperative functions of two CCCTC-binding factor (CTCF)-binding elements (CBE1 and CBE2) within IGCR1.
• To determine if CBE1 or CBE2 alone mediates IGCR1 functions in regulating IgH V(D)J recombination and repertoire diversity.

Main Methods:

• Generation of genetically modified mice with individually inactivated IGCR1 CBE1 or CBE2 using scrambled sequences.
• Analysis of IgH V(D)J recombination and V(H) gene usage in mutant mice.
• Comparison of the effects of individual CBE inactivation versus simultaneous inactivation of both CBEs.

Main Results:

• Individual inactivation of CBE1 or CBE2 resulted in only partial impairment of IGCR1 functions.
• Simultaneous inactivation of both CBE1 and CBE2 led to a significantly greater impairment of IGCR1 functions.
• These findings demonstrate a cooperative function between CBE1 and CBE2 in regulating IgH V(D)J recombination.

Conclusions:

• CBE1 and CBE2 within IGCR1 function cooperatively to fully regulate IgH V(D)J recombination and B-cell repertoire diversity.
• A model of orientation-specific looping is proposed to explain the synergistic functions of CBE1 and CBE2.