The “Dispensable” Portion of RAG2 Is Necessary for Efficient V-to-DJ Rearrangement during B and T Cell Development

Liang, Hong; Hsu, Lih Yun; Cado, Dragana; Cowell, Lindsay G.; Kelsoe, Garnett; Schlissel, Mark S.

doi:10.1016/s1074-7613(02)00448-x

Cited by 138 publications

(174 citation statements)

References 42 publications

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“…Second, unlike the core RAG proteins, RAG complexes containing full-length RAG2 cleave LMO2 more efficiently than Ttg-1, suggesting the C-terminal "noncore" portion of RAG2 influences the binding site specificity of the RAG complex (supplemental Fig. 2), as others have argued (31). We also acknowledge the possibility that distinct cRSSs may be similarly bound and cleaved by the RAG complex but support joining at different levels due to sequence variations (32).…”

Section: Discussionmentioning

confidence: 58%

V(D)J Recombinase Binding and Cleavage of Cryptic Recombination Signal Sequences Identified from Lymphoid Malignancies

Zhang

Swanson

2008

Journal of Biological Chemistry

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To understand the molecular basis for these large differences, we investigated the binding and cleavage of these cRSSs by the RAG1/2 proteins that initiate V(D)J recombination. We find that the RAG proteins comparably bind all cRSSs tested, albeit more poorly than a consensus RSS. We show that four cRSSs that support levels of V(D)J recombination above background levels in cell culture (LMO2, TAL1, Ttg-1, and SIL) are also cleaved by the RAG proteins in vitro with efficiencies ranging from 18 to 70% of a consensus RSS. Cleavage of LMO2 and Ttg-1 by the RAG proteins can also be detected in cell culture using ligation-mediated PCR. In contrast, Hox11 and SCL are nicked but not cleaved efficiently in vitro, and cleavage at other adventitious sites in plasmid substrates may also limit the ability to detect recombination activity at these cRSSs in cell culture.The antigen binding domains of immunoglobulins and T cell receptors are encoded in germ line arrays of V, D, and J gene segments that are assembled into functional variable exons by V(D)J recombination during lymphocyte development (1). The site of recombination is directed by a recombination signal sequence (RSS) 2 that flanks each receptor gene segment and consists of a conserved heptamer (consensus 5Ј-CACAGTG-3Ј) and nonamer (consensus 5Ј-ACAAAAACC-3Ј) separated by either 12 or 23 Ϯ 1 nucleotides of more highly varied sequence. V(D)J recombination can be conceptually divided into two phases, a cleavage phase and a joining phase (2). In the cleavage phase, the lymphoid cell-specific RAG1 and RAG2 (recombination activating gene-1 and -2, respectively) proteins assemble a multiprotein synaptic complex with two RSSs (generally one 12-RSS and one 23-RSS) and introduce a DNA double strand break at each RSS between the heptamer and adjacent coding segment via a nick-hairpin mechanism to yield a blunt 5Ј-phosphorylated signal end and a coding end terminating in a covalently sealed DNA hairpin structure. In the joining phase, the signal ends are generally joined precisely to form signal joints, and the coding ends are processed and joined to form coding joints that typically contain nucleotide additions or deletions at the junction. These processes are normally mediated by components of the nonhomologous end-joining DNA repair pathway.

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Section: Discussionmentioning

confidence: 58%

V(D)J Recombinase Binding and Cleavage of Cryptic Recombination Signal Sequences Identified from Lymphoid Malignancies

Zhang

Swanson

2008

Journal of Biological Chemistry

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“…To determine whether stable μH mRNA interferes with VDJ recombination by directly inhibiting the RAG recombinase, we transduced sorted pro-B cells from the bone marrow of homozygous Ter5 High and Ter5 Low mice with a VDJ recombination plasmid that contains an inverted EGFP gene. The EGFP cassette is flanked by recombination signal sequences (RSS) and can be activated upon RAG-mediated reversion (35). RAG activities in pro-B cells from both wild-type and Ter5 High mice were similar (Fig.…”

Section: Resultsmentioning

confidence: 95%

“…3B) and two fifths of that in Ter3 heterozygous pre-B cells. VDJ recombination at the TCR-β locus occurs in the DN III population (pro-T cell stage), and impaired recombination should increase this population (35). However, this and the other thymic cell populations were unaltered (Fig.…”

Section: Resultsmentioning

confidence: 99%

Pro-B cells sense productive immunoglobulin heavy chain rearrangement irrespective of polypeptide production

Lutz

Heideman

Roth

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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B-lymphocyte development is dictated by the protein products of functionally rearranged Ig heavy (H) and light (L) chain genes. Ig rearrangement begins in pro-B cells at the IgH locus. If pro-B cells generate a productive allele, they assemble a pre-B cell receptor complex, which signals their differentiation into pre-B cells and their clonal expansion. Pre-B cell receptor signals are also thought to contribute to allelic exclusion by preventing further IgH rearrangements. Here we show in two independent mouse models that the accumulation of a stabilized μH mRNA that does not encode μH chain protein specifically impairs pro-B cell differentiation and reduces the frequency of rearranged IgH genes in a dose-dependent manner. Because noncoding IgH mRNA is usually rapidly degraded by the nonsense-mediated mRNA decay machinery, we propose that the difference in mRNA stability allows pro-B cells to distinguish between productive and nonproductive Ig gene rearrangements and that μH mRNA may thus contribute to efficient H chain allelic exclusion.

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“…Mutation of the conserved tryptophan residue W453 within the PhD finger domain of RAG2 abrogates RAG2 binding to H3K4me3 and impairs V(D)J recombination of chromosomal and extra-chromosomal substrates (Liu et al, 2007;Matthews et al, 2007). However, removal of the entire RAG2 noncore region, including the PhD domain, only leads to a partial impairment of V(D)J recombination (Liang et al, 2002). These seemingly contradicting data have been suggested to reflect the presence of an inhibitory function within the noncore region of RAG-2, which is relieved upon binding to H3K4me3, or can be circumvented by deleting the entire noncore region (Liu et al, 2007;Matthews et al, 2007).…”

Section: Chromatin Modificationsmentioning

confidence: 99%

Chapter 1 Cis‐Regulatory Elements and Epigenetic Changes Control Genomic Rearrangements of the IgH Locus

Perlot

Alt

2008

Advances in Immunology

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Immunoglobulin variable region exons are assembled from discontinuous variable (V), diversity (D), and joining (J) segments by the process of V(D)J recombination. V(D)J rearrangements of the immunoglobulin heavy chain (IgH) locus are tightly controlled in a tissue-specific, ordered and allele-specific manner by regulating accessibility of V, D, and J segments to the recombination activating gene proteins which are the specific components of the V(D)J recombinase. In this review we discuss recent advances and established models brought forward to explain the mechanisms underlying accessibility control of V(D)J recombination, including research on germline transcripts, spatial organization, and chromatin modifications of the immunoglobulin heavy chain (IgH) locus. Furthermore, we review the functions of well-described and potential new cis-regulatory elements with regard to processes such as V(D)J recombination, allelic exclusion, and IgH class switch recombination.

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The “Dispensable” Portion of RAG2 Is Necessary for Efficient V-to-DJ Rearrangement during B and T Cell Development

Cited by 138 publications

References 42 publications

V(D)J Recombinase Binding and Cleavage of Cryptic Recombination Signal Sequences Identified from Lymphoid Malignancies

V(D)J Recombinase Binding and Cleavage of Cryptic Recombination Signal Sequences Identified from Lymphoid Malignancies

Pro-B cells sense productive immunoglobulin heavy chain rearrangement irrespective of polypeptide production

Chapter 1 Cis‐Regulatory Elements and Epigenetic Changes Control Genomic Rearrangements of the IgH Locus

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