Interdependence of N Nucleotide Addition and Recombination Site Choice in V(D)J Rearrangement

Kepler, Thomas B.; Borrero, Michelle; Rugerio, Beth; McCray, Suzanne; Clarke, Stephen H.

doi:10.4049/jimmunol.172.2.1311

Cited by 22 publications

(36 citation statements)

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“…The Ig V region repertoires differ between the perinatal and adult periods of life, with the former preferentially expressing D-proximal V H genes (1,69), showing distinct recombination sites (41), and, more importantly, lacking the diversifying potential of the N-nucleotide-producing TdT polymerase (8,27,32,44,55,57). In the absence of TdT activity, the usage of germ line SSH in the recombination between most coding gene partners is predominant.…”

Section: Discussionmentioning

confidence: 99%

A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

Gozalbo-López

Andrade

Terrados

et al. 2009

Molecular and Cellular Biology

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The molecular complexes involved in the nonhomologous end-joining process that resolves recombinationactivating gene (RAG)-induced double-strand breaks and results in V(D)J gene rearrangements vary during mammalian ontogeny. In the mouse, the first immunoglobulin gene rearrangements emerge during midgestation periods, but their repertoires have not been analyzed in detail. We decided to study the postgastrulation DJ H joints and compare them with those present in later life. The embryo DJ H joints differed from those observed in perinatal life by the presence of short stretches of nontemplated (N) nucleotides. Whereas most adult N nucleotides are introduced by terminal deoxynucleotidyl transferase (TdT), the embryo N nucleotides were due to the activity of the homologous DNA polymerase (Pol), which was widely expressed in the early ontogeny, as shown by analysis of Pol ؊/؊ embryos. Based on its DNA-dependent polymerization ability, which TdT lacks, Pol also filled in small sequence gaps at the coding ends and contributed to the ligation of highly processed ends, frequently found in the embryo, by pairing to internal microhomology sites. These findings show that Pol participates in the repair of early-embryo, RAG-induced double-strand breaks and subsequently may contribute to preserve the genomic stability and cellular homeostasis of lymphohematopoietic precursors during development.The adaptive immune system is characterized by the great diversity of its antigen receptors, which result from the activities of enzymatic complexes that cut and paste the genomic DNA of antigen receptor loci. The nonhomologous end-joining (NHEJ) machinery is then recruited to repair the doublestrand DNA breaks (DSBs) inflicted by the products of the recombination-activating genes (RAGs) (45,65). Within B cells, each immunoglobulin (Ig) receptor represents a singular shuffling of two heavy (H) and two light (L) chains, which are derived from the recombination of V, D, and J gene segments of the IgH locus and of V and J for IgL (71). Besides these combinatorial possibilities, most Ig variability derives from extensive processing of the coding ends, including exonucleolytic trimming of DNA ends, together with the addition of palindromic (P) nucleotides templated by the adjacent germ line sequence and of nontemplated (N) nucleotides secondary to the activity of the terminal deoxynucleotidyl transferase (TdT), a lymphoid-specific member of family X of DNA polymerases (reviewed in reference 56). During B-lineage differentiation, IgH rearrangements occur before those of the IgL locus, and D-to-J H rearrangements precede V-to-DJ H rearrangements (62). DJ H joints are formed in any of the three open reading frames (ORFs). ORF1 is predominantly used in mature Igs, ORF2 is transcribed as a D protein that provides negative signals to the B-cell precursors, and ORF3 frequently leads to stop codons (32,33,37). Germ line V, D, and J gene segments display short stretches of mutually homologous nucleotides (SSH), which are frequently used in gene rea...

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Section: Discussionmentioning

confidence: 99%

A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

Gozalbo-López

Andrade

Terrados

et al. 2009

Molecular and Cellular Biology

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“…The probabilities that N nucleotide addition could give rise to apparent D segments were determined by first identifying all unique 6-, 7-, 8-, 9-, 10-, 11-, and 12-nt sequences that can be produced from known D gene segments, and determining their probabilities using probabilities of N addition based upon the known TdT nucleotide preferences (15), as follows: p(A) ϭ 0.15, p(T) ϭ 0.15, p(G) ϭ 0.6, and p(C) ϭ 0.1. Calculations, which are more fully described below, included the equal likelihood that G nucleotides in N regions could arise from G addition to the sense strand, or from C addition to the antisense strand (20). The analysis was performed for complete matches, as well as for sequences with one or two mismatches.…”

Section: Partitioning Of Rearranged H Chain V Genesmentioning

confidence: 99%

“…To analyze whether putative N regions carried the G/C features of N regions, including high G or C content, and homogeneity of either G or C (20), the probability of the particular frequencies of G, C, and A/T nucleotides within the N regions was determined as follows:…”

Section: Analysis Of the Gc Content Of Putative N Regionsmentioning

confidence: 99%

“…The probabilities of an A or T insertion ( p W ) was set as 0.3, the probability of an insertion of G ( p G ) as 0.6, and the probability of C insertion ( p C ) as 0.1 as determined by Basu et al (15). The equation assumes that concatenation of strands does not occur (20), and allows for the fact that, for example, C additions may appear in the sense strand through addition of G nucleotides to the antisense strand.…”

Section: Analysis Of the Gc Content Of Putative N Regionsmentioning

confidence: 99%

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Partitioning of Rearranged Ig Genes by Mutation Analysis Demonstrates D-D Fusion and V Gene Replacement in the Expressed Human Repertoire

Collins

Ikutani

Puiu

et al. 2004

The Journal of Immunology

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The accurate partitioning of Ig H chain VHDJH junctions and L chain VLJL junctions is problematic. We have developed a statistical approach for the partitioning of such sequences, by analyzing the distribution of point mutations between a determined V gene segment and putative Ig regions. The establishment of objective criteria for the partitioning of sequences between VH, D, and JH gene segments has allowed us to more carefully analyze intervening putative nontemplated (N) nucleotides. An analysis of 225 IgM H chain sequences, with five or fewer V mutations, led to the alignment of 199 sequences. Only 5.0% of sequences lacked N nucleotides at the VHD junction (N1), and 10.6% at the DJH junction (N2). Long N regions (>9 nt) were seen in 20.6% of N1 regions and 17.1% of N2 regions. Using a statistical analysis based upon known features of N addition, and mutation analysis, two of these N regions aligned with D gene segments, and a third aligned with an inverted D gene segment. Nine additional sequences included possible alignments with a second D segment. Four of the remaining 40 long N1 regions included 5′ sequences having six or more matches to V gene end motifs, which may be the result of V gene replacement. Such sequences were not seen in long N2 regions. The long N regions frequently seen in the expressed repertoire of human Ig gene rearrangements can therefore only partly be explained by V gene replacement and D-D fusion.

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“…Recombination events in the heavy chain are more complex due to the presence of an additional D segment. One of the mechanisms of increasing diver sity is incorporation of n nucleotides (nontem plated nucleotides) at the gene segment joint site [4]. Later on, the DNA sequences of variable (V) domains are affected by the influence of somatic hypermutations taking place during the affinity maturation of B cells.…”

Section: In Silico Analysis Of the Structure Of Variable Domains Of Mmentioning

confidence: 99%

In silico analysis of the structure of variable domains of mouse single-chain antibodies specific to the human recombinant interferon β1b

2009

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Interdependence of N Nucleotide Addition and Recombination Site Choice in V(D)J Rearrangement

Abstract: The third author's last name was misspelled. The correct spelling is Ruggiero.

Cited by 22 publications

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A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

Partitioning of Rearranged Ig Genes by Mutation Analysis Demonstrates D-D Fusion and V Gene Replacement in the Expressed Human Repertoire

In silico analysis of the structure of variable domains of mouse single-chain antibodies specific to the human recombinant interferon β1b

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Interdependence of N Nucleotide Addition and Recombination Site Choice in V(D)J Rearrangement

Abstract: The third author's last name was misspelled. The correct spelling is Ruggiero.

Cited by 22 publications

References 0 publications

A Role for DNA Polymerase μ in the Emerging DJH Rearrangements of the Postgastrulation Mouse Embryo

A Role for DNA Polymerase μ in the Emerging DJH Rearrangements of the Postgastrulation Mouse Embryo

Partitioning of Rearranged Ig Genes by Mutation Analysis Demonstrates D-D Fusion and V Gene Replacement in the Expressed Human Repertoire

In silico analysis of the structure of variable domains of mouse single-chain antibodies specific to the human recombinant interferon β1b

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A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo

A Role for DNA Polymerase μ in the Emerging DJ_H Rearrangements of the Postgastrulation Mouse Embryo