Detection of immunoglobulin kappa light-chain gene rearrangement patterns by Southern blot analysis.

ABSTRACT

Immunoglobulin light-chain (IgL) gene rearrangements occur in a sequential order during normal B-cell differentiation with Ig kappa gene rearrangements prior to Ig lambda gene rearrangements. Therefore, Ig kappa producing B-cells usually retain Ig lambda genes in germline configuration, whereas the Ig kappa genes are generally deleted on one or both alleles in most Ig lambda producing B-cells. The deletion processes in the Ig kappa locus are mediated via rearrangement of the kappa deleting element (Kde), which is located approximately 24 kb downstream of the constant (C) kappa gene segment. Kde rearrangements can delete the C kappa region (including the Ig kappa enhancer) or the complete joining (J) kappa-C kappa region via rearrangements to a heptamer recombination signal sequence in the J kappa-C kappa intron (intron RSS), or via rearrangement to a variable (V) kappa gene segment, respectively. To improve the Southern blot detection of clonal Ig kappa gene rearrangements and deletions in B-lineage malignancies, we developed a new set of optimal J kappa, C kappa, and Kde probes, and made a detailed restriction map of the J kappa, C kappa, and Kde region. Extensive Southern blot studies revealed that rearrangements in the J kappa gene region are optimally detectable by use of a J kappa probe in combination with at least two appropriate restriction enzymes, i.e. BamHI, BglII, EcoRI, HindIII, and/or SacI. J kappa gene rearrangements are also detectable with the C kappa probe in BglII and BamHI digests, if no deletion of the C kappa region has occurred. The two different types of Kde-mediated J kappa and/or C kappa gene deletions are easily detectable with the Kde probe in BglII, HindIII and/or EcoRI digests. This is in contrast to the inaccurate information obtained with the J kappa and C kappa probes, because these probes can detect deletions only in the form of decreased densities of J kappa and/or C kappa germline bands in the absence of rearranged bands. Our detailed analysis of 217 B-lineage leukemias revealed that 62% (69/111) of precursor B-cell acute lymphoblastic leukemias had rearranged and/or deleted Ig kappa genes. All 53 Ig lambda+ chronic B-cell leukemias contained Ig kappa gene deletions; in 75% this concerned biallelic J kappa and/or C kappa gene deletions. Virtually all Ig kappa gene deletions appeared to be mediated via Kde rearrangements, while only 1.5% of the Ig kappa gene deletions were mediated via an alternative deletion mechanism which involved the J kappa region.