RAG1-mediated ubiquitylation of histone H3 is required for chromosomal V(D)J recombination

Deng, Zimu; Liu, Haifeng; Liu, Xiaolong

doi:10.1038/cr.2015.1

Cited by 31 publications

(47 citation statements)

References 37 publications

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“…This agrees with a recent modeling analysis that predicted that the RAG complex should localize efficiently to chromatin rich in H3K4me3 (Askary et al, 2014). Finally, given the ability of RAG1 to bind and ubiquitinate histone H3 (Deng et al, 2015; Grazini et al, 2010; Jones et al, 2011), it is plausible that direct RAG1-histone interactions contribute to the observed RAG1 binding pattern.…”

Section: Discussionmentioning

confidence: 99%

RAG Represents a Widespread Threat to the Lymphocyte Genome

Teng

Maman

Resch

et al. 2015

Cell

112

143

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SUMMARY The RAG1 endonuclease, together with its cofactor RAG2, is essential for V(D)J recombination but is a potent threat to genome stability. The sources of RAG1 mis-targeting and the mechanisms that have evolved to suppress it are poorly understood. Here, we report that RAG1 associates with chromatin at thousands of active promoters and enhancers in the genome of developing lymphocytes. The mouse and human genomes appear to have responded by reducing the abundance of “cryptic” recombination signals near RAG1 binding sites. This depletion operates specifically on the RSS heptamer, whereas nonamers are enriched at RAG1 binding sites. Reversing this RAG-driven depletion of cleavage sites by insertion of strong recombination signals creates an ectopic hub of RAG-mediated V(D)J recombination and chromosomal translocations. Our findings delineate rules governing RAG binding in the genome, identify areas at risk of RAG-mediated damage, and highlight the evolutionary struggle to accommodate programmed DNA damage in developing lymphocytes.

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Section: Discussionmentioning

confidence: 99%

RAG Represents a Widespread Threat to the Lymphocyte Genome

Teng

Maman

Resch

et al. 2015

Cell

112

143

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“…The non-core N-terminal region of RAG1 contains a series of basic motifs, among which the basic IIa domain (BIIa; amino acids 219–225) allows interaction with karyopherin subunit α1 (KPNA1; also known as importin subunit α5), which determines RAG1 subnuclear localization 52 . Following the basic motifs, a C3HC4 RING finger domain and an adjacent zinc finger motif form a single domain that coordinates zinc ions 50 and has histone H3 ubiquitin ligase activity, which is required for a normal level of chromosomal V(D)J recombination 53 . It has been proposed that the interaction of RAG1 with unubiquitylated H3 restrains RAG1 catalytic activity and that H3 ubiquitylation would release RAG1 and allow catalytic function 53 .…”

Section: Molecular Pathology Of Rag Deficiencymentioning

confidence: 99%

“…Following the basic motifs, a C3HC4 RING finger domain and an adjacent zinc finger motif form a single domain that coordinates zinc ions 50 and has histone H3 ubiquitin ligase activity, which is required for a normal level of chromosomal V(D)J recombination 53 . It has been proposed that the interaction of RAG1 with unubiquitylated H3 restrains RAG1 catalytic activity and that H3 ubiquitylation would release RAG1 and allow catalytic function 53 .…”

Section: Molecular Pathology Of Rag Deficiencymentioning

confidence: 99%

Human RAG mutations: biochemistry and clinical implications

et al. 2016

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The recombination-activating gene 1 (RAG1) and RAG2 proteins initiate the V(D)J recombination process, which ultimately enables the generation of T cells and B cells with a diversified repertoire of antigen-specific receptors. Mutations of the RAG genes in humans are associated with a broad spectrum of clinical phenotypes, ranging from severe combined immunodeficiency to autoimmunity. Recently, novel insights into the phenotypic diversity of this disease have been provided by resolving the crystal structure of the RAG complex, by developing novel assays to test recombination activity of the mutant RAG proteins and by characterizing the molecular and cellular basis of immune dysregulation in patients with RAG deficiency.

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“…Perhaps also relevant to the interpretation of RAG1 binding patterns is the finding that RAG1 can interact with histone 3 (46,47). This emphasizes the point that our ChIP assay does not distinguish between interactions of the RAG proteins directly with DNA and interactions with the protein components of chromatin.…”

Section: Discussionmentioning

confidence: 91%

Recruitment of RAG1 and RAG2 to Chromatinized DNA during V(D)J Recombination

Shetty

Schatz

2015

Molecular and Cellular Biology

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b V(D)J recombination is initiated by the binding of the RAG1 and RAG2 proteins to recombination signal sequences (RSSs) that consist of conserved heptamer and nonamer sequences separated by a spacer of either 12 or 23 bp. Here, we used RAG-inducible pro-B v-Abl cell lines in conjunction with chromatin immunoprecipitation to better understand the protein and RSS requirements for RAG recruitment to chromatin. Using a catalytic mutant form of RAG1 to prevent recombination, we did not observe cooperation between RAG1 and RAG2 in their recruitment to endogenous J gene segments over a 48-h time course. Using retroviral recombination substrates, we found that RAG1 was recruited inefficiently to substrates lacking an RSS or containing a single RSS, better to substrates with two 12-bp RSSs (12RSSs) or two 23-bp RSSs (23RSSs), and more efficiently to a substrate with a 12/23RSS pair. RSS mutagenesis demonstrated a major role for the nonamer element in RAG1 binding, and correspondingly, a cryptic RSS consisting of a repeat of CA dinucleotides, which poorly re-creates the nonamer, was ineffective in recruiting RAG1. Our findings suggest that 12RSS-23RSS cooperation (the "12/23 rule") is important not only for regulating RAG-mediated DNA cleavage but also for the efficiency of RAG recruitment to chromatin. T he adaptive immune system relies on V(D)J recombination to assemble variable (V), diversity (D), and joining (J) antigen receptor gene segments in generating a diverse repertoire of immunoglobulins and T cell receptors. The process is initiated by the recombination-activating gene (RAG) recombinase, a protein complex consisting of the enzymes encoded by RAG1 and RAG2 (1, 2). RAG1 is 1,040 amino acids long, with a core region spanning amino acids 384 to 1008 that contains recombination signal sequence (RSS) and RAG2 binding activity as well as the catalytic residues responsible for DNA cleavage (3-7). RAG2 is 527 amino acids long, with a core region (amino acids 1 to 387) that interacts with and facilitates DNA binding and cleavage by RAG1 but which has no DNA binding activity on its own (3,4,8,9). The RAG2 noncore region contains a plant homeodomain (PHD) finger that plays an important role in chromatin binding through interactions with the histone H3 N-terminal tail when lysine 4 is trimethylated (H3K4me3) (10-12).The site of recombination is specified by the RSS that immediately flanks each V, D, and J gene segment (Fig. 1A). The RSS consists of a well-conserved 7-bp sequence called the heptamer (consensus sequence of 5=-CACAGTG-3=) and an AT-rich 9-bp sequence called the nonamer (consensus sequence of 5=-ACAAA AACC-3=) separated by a poorly conserved spacer whose length is either 12 or 23 bp (13-15) (Fig. 1B). An RSS with a spacer 12 bp long is defined as a 12RSS, while one with a spacer 23 bp long is defined as a 23RSS (13).The consensus RSS is the most efficiently recombined sequence and therefore has been used for most in vitro experiments; however, endogenous RSSs often deviate from the consensus at multiple hept...

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RAG1-mediated ubiquitylation of histone H3 is required for chromosomal V(D)J recombination

Cited by 31 publications

References 37 publications

RAG Represents a Widespread Threat to the Lymphocyte Genome

RAG Represents a Widespread Threat to the Lymphocyte Genome

Human RAG mutations: biochemistry and clinical implications

Recruitment of RAG1 and RAG2 to Chromatinized DNA during V(D)J Recombination

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