A functional analysis of the spacer of V(D)J recombination signal sequences.
Abstract
During lymphocyte development, V(D)J recombination assembles antigen receptor genes
from component V, D, and J gene segments. These gene segments are flanked by a recombination
signal sequence (RSS), which serves as the binding site for the recombination machinery.
The murine Jbeta2.6 gene segment is a recombinationally inactive pseudogene, but examination
of its RSS reveals no obvious reason for its failure to recombine. Mutagenesis of
the Jbeta2.6 RSS demonstrates that the sequences of the heptamer, nonamer, and spacer
are all important. Strikingly, changes solely in the spacer sequence can result in
dramatic differences in the level of recombination. The subsequent analysis of a library
of more than 4,000 spacer variants revealed that spacer residues of particular functional
importance are correlated with their degree of conservation. Biochemical assays indicate
distinct cooperation between the spacer and heptamer/nonamer along each step of the
reaction pathway. The results suggest that the spacer serves not only to ensure the
appropriate distance between the heptamer and nonamer but also regulates RSS activity
by providing additional RAG:RSS interaction surfaces. We conclude that while RSSs
are defined by a "digital" requirement for absolutely conserved nucleotides, the quality
of RSS function is determined in an "analog" manner by numerous complex interactions
between the RAG proteins and the less-well conserved nucleotides in the heptamer,
the nonamer, and, importantly, the spacer. Those modulatory effects are accurately
predicted by a new computational algorithm for "RSS information content." The interplay
between such binary and multiplicative modes of interactions provides a general model
for analyzing protein-DNA interactions in various biological systems.
Type
Journal articleSubject
AlgorithmsAnimals
Cell Line
Cloning, Molecular
Computational Biology
DNA
DNA, Intergenic
Humans
Lymphocytes
Mice
Models, Genetic
Models, Statistical
Mutagenesis
Oligonucleotides
Plasmids
Protein Binding
Protein Sorting Signals
Recombination, Genetic
Software
T-Lymphocytes
VDJ Recombinases
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https://hdl.handle.net/10161/11485Published Version (Please cite this version)
10.1371/journal.pbio.0000001Publication Info
Lee, Alfred Ian; Fugmann, Sebastian D; Cowell, Lindsay G; Ptaszek, Leon M; Kelsoe,
Garnett; & Schatz, David G (2003). A functional analysis of the spacer of V(D)J recombination signal sequences. PLoS Biol, 1(1). pp. E1. 10.1371/journal.pbio.0000001. Retrieved from https://hdl.handle.net/10161/11485.This is constructed from limited available data and may be imprecise. To cite this
article, please review & use the official citation provided by the journal.
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Show full item recordScholars@Duke
Lindsay Grey Cowell
Adjunct Assistant Professor in the Department of Biostatistics and Bioinformatics
Somatic Diversification of Lymphocyte Antigen Receptor Genes * V(D)J Recombination
* Somatic Hypermutation Biomedical Ontology * Ontological Representation of Cells
of Hematopoietic Lineage Biomedical Text Mining Logic-based Reasoning
Garnett H. Kelsoe
James B. Duke Distinguished Professor of Immunology
1. Lymphocyte development and antigen-driven diversification of immunoglobulin and
T cell antigen receptor genes. 2. The germinal center reaction and mechanisms for
clonal selection and self - tolerance. The origins of autoimmunity. 3. Interaction
of innate- and adaptive immunity and the role of inflammation in lymphoid organogenesis.
4. The role of secondary V(D)J gene rearrangment in lymphocyte development and malignancies.
5. Mathematical modeling of immune responses,
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