No evidence for the use of DIR, D–D fusions, chromosome 15 open reading frames or VHreplacement in the peripheral repertoire was found on application of an improved algorithm, JointML, to 6329 human immunoglobulin H rearrangements

Abstract

Antibody diversity is created by imprecise joining of the variability (V), diversity (D) and joining (J) gene segments of the heavy and light chain loci. Analysis of rearrangements is complicated by somatic hypermutations and uncertainty concerning the sources of gene segments and the precise way in which they recombine. It has been suggested that D genes with irregular recombination signal sequences (DIR) and chromosome 15 open reading frames (OR15) can replace conventional D genes, that two D genes or inverted D genes may be used and that the repertoire can be further diversified by heavy chain V gene (VH) replacement. Safe conclusions require large, well-defined sequence samples and algorithms minimizing stochastic assignment of segments. Two computer programs were developed for analysis of heavy chain joints. JointHMM is a profile hidden Markow model, while JointML is a maximum-likelihood-based method taking the lengths of the joint and the mutational status of the VH gene into account. The programs were applied to a set of 6329 clonally unrelated rearrangements. A conventional D gene was found in 80% of unmutated sequences and 64% of mutated sequences, while D-gene assignment was kept below 5% in artificial (randomly permutated) rearrangements. No evidence for the use of DIR, OR15, multiple D genes or VH replacements was found, while inverted D genes were used in less than 1 per thousand of the sequences. JointML was shown to have a higher predictive performance for D-gene assignment in mutated and unmutated sequences than four other publicly available programs. An online version 1.0 of JointML is available at http://www.cbs.dtu.dk/services/VDJsolver.