Impaired V(D)J Recombination and Lymphocyte Development in Core RAG1-expressing Mice

RAG1 and RAG2 are the lymphocyte-specific components of the V(D)J recombinase. In vitro analyses of RAG function have relied on soluble, highly truncated “core” RAG proteins. To identify potential functions for noncore regions and assess functionality of core RAG1 in vivo, we generated core RAG1 knockin (RAG1c/c) mice. Significant B and T cell numbers are generated in RAG1c/c mice, showing that core RAG1, despite missing ∼40% of the RAG1 sequence, retains significant in vivo function. However, lymphocyte development and the overall level of V(D)J recombination are impaired at the progenitor stage in RAG1c/c mice. Correspondingly, there are reduced numbers of peripheral RAG1c/c B and T lymphocytes. Whereas normal B lymphocytes undergo rearrangement of both JH loci, substantial levels of germline JH loci persist in mature B cells of RAG1c/c mice, demonstrating that DJH rearrangement on both IgH alleles is not required for developmental progression to the stage of VH to DJH recombination. Whereas VH to DJH rearrangements occur, albeit at reduced levels, on the nonselected alleles of RAG1c/c B cells that have undergone D to JH rearrangements, we do not detect VH to DH rearrangements in RAG1c/c B cells that retain germline JH alleles. We discuss the potential implications of these findings for noncore RAG1 functions and for the ordered assembly of VH, DH, and JH segments

Recombination-activating gene proteins: more regulation, please

During V(D)J recombination, developing B and T cells assemble gene segments in order to create the variable regions of immunoglobulin and T cell receptors required by our adaptive immune response. The chemistry of this recombination pathway requires a specific nuclease, and a more general repair pathway for double strand breaks. The nuclease activity is provided by a complex of the RAG1 and RAG2 proteins. In fact, RAG1 and RAG2 probably coordinate many steps involving the coding and signal DNA sequences. Studies using deletion and truncation mutants of the RAG proteins demonstrate that each of these contain a functional core region, representing about two-thirds of each polypeptide, While the core regions are sufficient to catalyze recombination in test systems, the full-length proteins seem to show more complicated behaviors in vivo. A plausible explanation is that regions outside the core help in the proper regulation of recombination. The non-core region of RAG1 has just been found to contain a ubiquitin ligase. Regulatory functions may contribute to autoregulation of the proteins involved, fidelity of the reaction, protection of the cell from translocations, coordination of recombination with the cell-cycle and possibly modification of the chromatin structure of target DNA

Educational Case: Benign Papilloma of the Breast

The following fictional case is intended as a learning tool within the Pathology Competencies for Medical Education (PCME), a set of national standards for teaching pathology. These are divided into three basic competencies: Disease Mechanisms and Processes, Organ System Pathology, and Diagnostic Medicine and Therapeutic Pathology. For additional information, and a full list of learning objectives for all three competencies, see http://journals.sagepub.com/doi/10.1177/2374289517715040

The RAG1 N-terminal domain is an E3 ubiquitin ligase

RAG1 and RAG2 initiate V(D)J recombination, which is the assembly of immunoglobulin and T cell receptor genes. The N-terminal region of RAG1 can be deleted, leaving an enzymatic “core” able to catalyze the complete reaction. Here we report that the N-terminal portion of RAG1 has a distinct enzymatic role separate from the rest of the protein. It acts as an E3 ligase in the ubiquitylation of a test substrate and formation of polyubiquitin chains in vitro. This finding suggests a new way in which V(D)J recombination can be regulated and coupled to other aspects of cell physiology