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Chromosomal translocations involving the immunoglobulin (Ig) receptor loci usually disrupt and silence these loci. On the basis of observations in follicular lymphoma (FL) with downstream Ig heavy chain (IGH) class switch recombination (CSR), we hypothesized that downstream CSR-mediated chromosomal translocations would leave the V(D)J-Cmu transcription unit intact, thereby still allowing IgM expression from the IGH allele involved in the translocation. To test this hypothesis, we analyzed biallelic IGH translocations in the IgM-expressing cell line Z-138 by interphase FISH, DNA fiber-FISH, long-distance vectorette PCR, and DNA sequencing. One IGH allele was involved in a t(11;14), showing a break in the JH region that juxtaposed the El,L enhancer and the 3' Calpha enhancers to the cyclin D1 gene. The other IGH allele contained a t(8; 14) breakpoint involving the 3' end of a Sgamma region, whereas the reciprocal breakpoint at 8q24 was approximately 40 kb centromeric of MYC. Molecular analysis showed that this IGH allele harbored a normal V(D)J-Cmu complex, which is responsible for IgM expression. These data show that chromosomal breakpoints such as the t(8; 14) can occur in downstream IGH constant regions and do not necessarily interfere with Ig expression. (C) 2004 Wiley-Liss, Inc.

Most chromosomal t(8;14) translocations in sporadic Burkitt lymphomas (BL) are mediated by immunoglobulin class switch, recombination (CSR), yet all tumors express IgM, suggesting an incomplete or exclusively monoallelic CSR event. We studied the exact configuration of both the nontranslocated IGH allele and the MYG/IGH breakpoint by applying a combination of low- and high-resolution methods (interphase FISH, DNA fiber FISH, long-distance PCR, and Southern blotting) on 16 BL. IGH class switch events involving the nontranslocated IGH allele were not observed. Thirteen cases had MYG/IGH breakpoints in or nearby IGH switch (S) sites, including five at S mu, three at S gamma and five at S alpha. All eight translocations with a breakpoint at S gamma or S alpha were perfectly reciprocal, without deletion of C mu-C delta or other CH elements. Internal S mu deletions claimed to be a marker for CSR activity and implicated in stabilization of IgM expression were found in BL but did riot correlate with downstream translocation events. This study shows that switch breakpoints in sporadic BL are exclusively resolved by a noncanonical recombination mechanism involving only one switch region. (c) 2006 Wiley-Liss, Inc.

Monoallelic expression of immunoglobulin genes is epigenetically regulated and maintained. Asynchronous replication of the IGH alleles has been implicated in allelic exclusion. Usually, the functional IGH allele replicates early in the S-phase of the cell cycle whereas the non-functional allele replicates late. Previously, the intronic enhancer region (Eμ) and the 3′ Cα enhancer region have been designated as putative replication initiation sites. Activity of these replication origins are likely to be involved in regulation of asynchronous replication. By use of interphase and DNA fiber FISH we have performed a detailed analysis of the configuration of the t(8;14) chromosomal translocation in Burkitt’s lymphoma patients and cell lines and showed that the breakpoints in all studied cases were perfectly reciprocal without loss of IGH genomic material. An important implication is that in patients and cell lines harboring a breakpoint in a downstream switch region (Sγ or Sα) the Eμ and the 3′ Cα enhancer region are physically separated from each other, whereas in patients and cell lines with the IGH breakpoint located in the JH-region both enhancer regions remain on the der(14) chromosome. We therefore studied the IGH replication timing in Burkitt’s lymphoma cell lines harboring different IGH breakpoints by use of BrdU-FISH and interphase FISH on sorted cell cycle fractions. In two cell lines with JH-region breakpoints (Jiyoye and DG-75) the t(8;14) was invariably targeted to the late replicating IGH allele. In contrast, in all three cell lines with switch-region breakpoints (CA-46, Namalwa, BL-65) asynchronous replication of the IGH alleles was lost. As the position of the breakpoint in the MYC locus at 8q24 differed substantially between these cell lines (DG-75 and CA-46: intron 1 of the c-myc gene; Jiyoye, Namalwa, and BL-65: 100 kb to >500 kb centromeric from c-myc) it is unlikely that the position of the 8q24 breakpoint is of crucial influence on IGH replication timing. We speculate that the Eμ and the 3′ Cα enhancer region regulate the IGH replication timing in a cis-acting manner as physical separation of both enhancer regions results in loss of asynchronous replication.