Immunoglobulin Heavy Chain Switch Regions Research Articles

CtIP-mediated DNA resection is dispensable for IgH class switch recombination by alternative end-joining

To generate antibodies with different effector functions, B cells undergo Immunoglobulin Heavy Chain (IgH) class switch recombination (CSR). The ligation step of CSR is usually mediated by the classical nonhomologous end-joining (cNHEJ) pathway. In cNHEJ-deficient cells, a remarkable ∼25% of CSR can be achieved by the alternative end-joining (Alt-EJ) pathway that preferentially uses microhomology (MH) at the junctions. While A-EJ-mediated repair of endonuclease-generated breaks requires DNA end resection, we show that CtIP-mediated DNA end resection is dispensable for A-EJ-mediated CSR using cNHEJ-deficient B cells. High-throughput sequencing analyses revealed that loss of ATM/ATR phosphorylation of CtIP at T855 or ATM kinase inhibition suppresses resection without altering the MH pattern of the A-EJ-mediated switch junctions. Moreover, we found that ATM kinase promotes Alt-EJ-mediated CSR by suppressing interchromosomal translocations independent of end resection. Finally, temporal analyses reveal that MHs are enriched in early internal deletions even in cNHEJ-proficient B cells. Thus, we propose that repetitive IgH switch regions represent favored substrates for MH-mediated end-joining contributing to the robustness and resection independence of A-EJ-mediated CSR.

DNA Replication Origins in Immunoglobulin Switch Regions Regulate Class Switch Recombination in an R-Loop-Dependent Manner

Class switch recombination (CSR) at the immunoglobulin heavy chain (IgH) locus generates antibody isotypes. CSR depends on double-strand breaks (DSBs) induced by activation-induced cytidine deaminase (AID). Although DSB formation and repair machineries are active in G1 phase, efficient CSR is dependent on cell proliferation and S phase entry; however, the underlying mechanisms are obscure. Here, we show that efficient CSR requires the replicative helicase, the Mcm complex. Mcm proteins are enriched at IgH switch regions during CSR, leading to assembly of facultative replication origins that require Mcm helicase function for productive CSR. Assembly of CSR-associated origins is facilitated by R loops and promotes the physical proximity (synapsis) of recombining switch regions, which is reduced by R loop inhibition or Mcm complex depletion. Thus, R loops contribute to replication origin specification that promotes DSB resolution in CSR. This suggests a mechanism for the dependence of CSR on S phase and cell division.

Erroneous class switching and false VDJ recombination: Molecular dissection of t(8;14)/MYC-IGH translocations in Burkitt-type lymphoblastic leukemia/B-cell lymphoma

The chromosomal translocation t(8;14)(q24;q32) with juxtaposition of MYC to enhancer elements in the immunoglobulin heavy chain (IGH) gene locus is the genetic hallmark of the majority of Burkitt lymphoma and a subset of Diffuse large B-cell lymphoma patients. Around 3% of adult B-lineage acute lymphoblastic leukemia (ALL) patients show this aberration. Flow cytometry mostly reveals a “mature B-ALL” or “Burkitt-type” ALL immunophenotype. Using long-distance PCR for t(8;14)/MYC-IGH fusion, we investigated bone marrow, peripheral blood and a few other samples with suspected Burkitt-ALL or mature B-ALL and identified 133 MYC-IGH-positive cases. The location of the chromosomal breaks in the IGH joining and the 8 different switch regions was determined using a set of long-distance PCRs. The chromosomal breakpoints with the adjacent MYC regions on 8q24 were characterized by direct sequencing in 49 cases. The distribution of chromosomal breaks among the IGH joining and switch regions was the following: JH 23.3%, M 21.8%, G1 15.0%, G2 7.5%, G3 3.8%, G4 4.5%, A1 12.8%, A2 3.8%, E 7.5%. Two breakpoint clusters near MYC were delineated. There was no clear correlation between the degree of somatic hypermutation and the chromosomal break locations. Epstein Barr virus was detected in 5 cases (4%). This detailed and extensive molecular analysis illustrates the molecular complexity of the MYC-IGH translocations and the detected distribution of breakpoints provides additional evidence that this translocation results from failed switch and VDJ recombinations. This study may serve as a model for the analysis of other IGH translocations in B-cell lymphoma.

Structure and Consequences of IGH Switch Breakpoints in Burkitt Lymphoma

The t(8;14) MYC/IGH breakpoint is the hallmark translocation of human Burkitt lymphoma (BL). The translocation breakpoint most often involves the immunoglobulin heavy-chain switch regions and is thought to be brought about by an aberrant class switch recombination (CSR) event. During CSR in normal germinal center B cells, DNA double-stranded breaks are introduced in Smu and one of the downstream switch regions (Sgamma, Salpha, or Sepsilon) that are juxtaposed and ligated to form the switch junction, with deletion of the intervening DNA. In contrast, aberrant switch recombination in BL exclusively involves only one switch region, resulting in a perfect reciprocal translocation. A functional consequence of this type of translocation is that IgM expression from the chromosome affected by the translocation is not necessarily disrupted.

The Human RecQ Helicases, BLM and RECQ1, Display Distinct DNA Substrate Specificities

RecQ helicases maintain chromosome stability by resolving a number of highly specific DNA structures that would otherwise impede the correct transmission of genetic information. Previous studies have shown that two human RecQ helicases, BLM and WRN, have very similar substrate specificities and preferentially unwind noncanonical DNA structures, such as synthetic Holliday junctions and G-quadruplex DNA. Here, we extend this analysis of BLM to include new substrates and have compared the substrate specificity of BLM with that of another human RecQ helicase, RECQ1. Our findings show that RECQ1 has a distinct substrate specificity compared with BLM. In particular, RECQ1 cannot unwind G-quadruplexes or RNA-DNA hybrid structures, even in the presence of the single-stranded binding protein, human replication protein A, that stimulates its DNA helicase activity. Moreover, RECQ1 cannot substitute for BLM in the regression of a model replication fork and is very inefficient in displacing plasmid D-loops lacking a 3'-tail. Conversely, RECQ1, but not BLM, is able to resolve immobile Holliday junction structures lacking an homologous core, even in the absence of human replication protein A. Mutagenesis studies show that the N-terminal region (residues 1-56) of RECQ1 is necessary both for protein oligomerization and for this Holliday junction disruption activity. These results suggest that the N-terminal domain or the higher order oligomer formation promoted by the N terminus is essential for the ability of RECQ1 to disrupt Holliday junctions. Collectively, our findings highlight several differences between the substrate specificities of RECQ1 and BLM (and by inference WRN) and suggest that these enzymes play nonoverlapping functions in cells.

TransCGH: Rapid Identification and High-Resolution Mapping of Balanced IgH Translocations in Archival DNA Using Custom Oligonucleotide Arrays.

Array based comparative genomic hybridization (CGH) has revolutionized the study of chromosomal imbalances but generally is incapable of detecting balanced genomic rearrangements like reciprocal translocations, which play central roles in the pathogenesis and diagnosis of lymphomas, leukemias and other tumors. The precise identification of immunoglobulin heavy chain (IgH) translocation partners, for example, is essential for the classification of B cell lymphomas and for predicting prognosis in plasma cell neoplasms like multiple myeloma. Using IgH translocations as a model for balanced genomic rearrangements, we have developed a simple modification of array CGH that we call translocation-CGH (transCGH) and that enables the rapid identification of IgH translocation partners and precise mapping of translocation-associated breakpoints to unprecedented resolution. To render IgH translocations detectable on CGH arrays, genomic DNA from test and reference samples is modified prior to array hybridization in an enzymatic linear amplification reaction that employs a single IgH joining (JH) or switch (Sμ/Sα/Sε) region primer, resulting in specific amplification of any fusion partner sequences that may be inserted (via translocation or other rearrangement) downstream of the IgH primer. Using a single tiling-density oligonucleotide array representing such common IgH partner loci as MYC, BCL2 and CCND1 (cyclin D1), transCGH successfully identified and mapped to ∼100bp resolution an assortment of known IgH fusion breakpoints in various cell lines and primary lymphomas, including JH-CCND1 breakpoints in MO2058 and Granta 519 cell lines (mantle cell lymphoma), a cytogenetically cryptic Sα-CCND1 fusion in U266 (myeloma), JH-MYC and Sμ-MYC breakpoints in MC116 and Raji (Burkitt lymphoma), and JH-BCL2 breakpoints in DHL16 (large cell lymphoma; minor cluster region) and in an archival case of follicular lymphoma (major breakpoint region). We then used transCGH to analyze 4 archival cases of mantle cell lymphoma and one t(11;14)-positive case of B cell prolymphocytic leukemia, all of which lacked PCR-detectable translocation breakpoints at the CCND1 major translocation cluster (MTC). Five novel CCND1 translocation breakpoints were identified and mapped to ∼100bp resolution, allowing the rapid design of patient-specific PCR primers for amplification, sequencing, and confirmation of the predicted breakpoints. One breakpoint mapped to within 500bp of the MTC, whereas the other 4 were scattered across a ∼150kb region flanking the MTC. To our knowledge, this represents the largest series of non-MTC mantle cell lymphoma breakpoint sequences reported to date. It also illustrates how transCGH can facilitate the rapid cloning of previously unidentified IgH translocation breakpoints dispersed over very large genomic regions. Because transCGH requires only genomic DNA and can simultaneously detect both balanced IgH translocations and genomic imbalances at ultra-high resolution on the same array, it may become a useful alternative to molecular cytogenetic methods (e.g. FISH) for clinical testing of B cell and plasma cell neoplasms. transCGH also will facilitate the development of highly sensitive breakpoint-specific PCR assays for detecting minimal residual disease. Finally, because the primer used in the linear amplification reaction is fully customizable, transCGH can readily be adapted to identify and map other balanced translocations (or more complex genomic fusions) that involve non-IgH loci, provided that one of the fusion partners is known.

Microfluidic Chips for Detecting the t(4;14) Translocation and Monitoring Disease during Treatment Using Reverse Transcriptase-Polymerase Chain Reaction Analysis of IgH-MMSET Hybrid Transcripts

Diagnosis platforms incorporating low-cost microfluidic chips enable sensitive, rapid, and accurate genetic analysis that could facilitate customized therapies tailored to match the vulnerabilities of any types of cancer. Using ex vivo cancer cells, we have detected the unique molecular signature and a chromosomal translocation in multiple myeloma. Multiple myeloma is characterized by IgH rearrangements and translocations that enable unequivocal identification of malignant cells, detected here with integrated microfluidic chips incorporating genetic amplification via reverse transcriptase-polymerase chain reaction and capillary electrophoresis. On microfluidic chips, we demonstrated accurate and versatile detection of molecular signatures in individual cancer cells, with value for monitoring response to therapy, detecting residual cancer cells that mediate relapse, and evaluating prognosis. Thus, testing for two clinically important molecular biomarkers, the IgH VDJ signature and hybrid transcripts signaling the t(4;14) chro-mosomal translocation, with predictive value in diagnosis, treatment decisions, and monitoring has been efficiently implemented on a miniaturized microfluidic system.

Treatment of Newly Diagnosed Multiple Myeloma Based on Mayo Stratification of Myeloma and Risk-Adapted Therapy (mSMART): Consensus Statement

Treatment of Newly Diagnosed Multiple Myeloma Based on Mayo Stratification of Myeloma and Risk-Adapted Therapy (mSMART): Consensus Statement

Bone Involvement in Multiple Myeloma Patients Carrying the T (4;14) Chromosomal Translocation.

Chromosomal translocations involving the immunoglobulin heavy chain switch region (IgH) are quite common in multiple myeloma (MM), and some of them can reliably predict disease outcome. In particular, t(4;14) chromosomal abnormality is one of the most adverse prognostic factors for response duration and survival after high dose therapy and autologous stem cell transplantation. Despite the dismal prognosis, however, in this subset of patients, bone involvement, as evaluated by spine MRI, is relatively infrequent, at variance to what has been observed in other MM subtypes according to TC classification. In the present study we aimed at further testing this hypothesis by analyzing the extent of whole bone involvement in patients showing t(4;14) chromosomal translocation as compared to negative patients. For this purpose, 50 newly diagnosed MM patients (32M, 18F, median age = 54 yrs) underwent evaluation of total skeletal X-ray, whole spine MRI and, at the same time, quantification of markers of bone resorption (urinary NTX, PYR and DPYR and serum crosslaps) and bone formation (bone alkaline phosphatase-BAP and osteocalcin) was performed. Using a real-time PCR assay to detect the presence of IgH/MMSET fusion gene as a surrogate marker for t(4;14), we found 15 patients carrying this chromosomal abnormality, 7 of whom (46%) were also positive for the deletion of chromosome 13, this abnormality was detected in 11/35 (31%) patients who proved negative for IgH/MMSET hybrid transcript. The two groups of patients did not differ significantly in terms of median age, distribution of M protein isotype and light chain, beta-2 microglobulin, bone marrow plasma cell infiltration and disease stage. Spinal MRI was negative in 3/15 (20%) t(4;14) positive patients as compared to 12% t(4;14) negative patients; skeletal involvement, however, was more pronounced in t(4;14) positive patients (median skeletal score = 6.24, as compared to 3.58 in t(4;14) negative cases, p= 0.00). These data were confirmed by the evaluation of bone resorption markers; specifically, serum crosslaps were significantly increased in patients with t(4;14) abnormality compared to negative individuals (7984 pmol/L±1682SE vs 5123pmol/L±783SE p=0.04). Conversely, no difference in bone formation markers was found in the two groups of patients. Our results indicate that, despite a spinal involvement at MRI that is comparable to what is observed in negative patients, individuals who are t(4;14) positive show a more pronounced bone resorption pattern, this in contrast to what has been reported so far, but in line with the aggressive features of the disease in these patients.

Molecular heterogeneity of sporadic adult Burkitt-type leukemia/lymphoma as revealed by PCR and cytogenetics: correlation with morphology, immunology and clinical features

Chromosomal translocations involving the MYC oncogene are a hallmark of Burkitt lymphoma but they are only found in a varying frequency in mature Burkitt-type acute lymphoblastic leukemia (B-ALL). We have investigated samples of 56 sporadic Burkitt leukemia/lymphoma patients for the translocations t(8;14)(q24;q32), t(2;8)(p11;q24) and t(8;22)(q24;q11). Long PCR was used for detecting the immunoglobulin heavy chain (IgH) translocation and cytogenetics and/or fluorescence in situ hybridization for detecting the 'variant' MYC translocations. A total of 29 samples (51.8%) were t(8;14)-positive by long PCR. Approximately one-third had a chromosomal breakpoint in the IgH joining region while the others had breakpoints in the IgH switch regions. Among them were two cases with a previously unreported MYC translocation into the IgE switch region. Long PCR was more reliable compared to conventional cytogenetics for detecting the t(8;14). Epstein-Barr virus was detected in high copy number in two (3.6%) t(8;14)-positive cases by real-time quantitative PCR. Human herpesvirus 8 was not detected in any case by nested PCR. A typical L3 or L3-compatible cytomorphology was highly predictive (>80%) but not specific of a MYC translocation. A total of 34 patients were treated according to the GMALL B-ALL therapy protocols and there was no significant difference in overall survival between patients with or without t(8;14).

Insertional events as well as translocations may arise during aberrant immunoglobulin switch recombination in a patient with multiple myeloma.

The majority of patients with multiple myeloma have translocations involving the immunoglobulin heavy chain switch regions on chromosome 14q32 and a promiscuous range of partner chromosomes. We describe a patient with an insertion of 132 bp of chromosome 22q12 sequence into the 5' region flanking S(mu) on chromosome 14q32. The 132 bp region from chromosome 22q12 contains the whole of exon 3 from a novel gene of unknown function in man. The significance of such insertional events remains unclear. The description of insertional events occurring as a result of abnormal switch recombination suggests that, in myeloma, dysregulation of oncogenes may occur by a mechanism other than chromosomal translocation.

Detection of aberrant isotype switch recombination in low-grade and high-grade gastric MALT lymphomas

Gastric mucosa-associated lymphoid tissue (MALT) lymphoma originates from reactive lymphocytic infiltrates during chronic gastritis, closely associated with Helicobacter pylori infection. MALT lymphomas may be either "low grade" or "high grade," and transformation from low grade to high grade can occur. To obtain information on the maturational state of MALT lymphoma cells, we investigated their ability to undergo isotype switch recombination, which together with immunoglobulin variable gene somatic mutation, contributes to normal B-cell maturation. Using specific probes for the immunoglobulin heavy-chain (IgH) switch regions, we found by Southern blot that 3 out of 5 low-grade cases and 2 out of 2 high-grade cases showed rearrangements within IgH switch regions, which appeared aberrant in 4 of the 5 cases. The cloning of two rearranged fragments from one low-grade and one high-grade case confirmed the aberrant nature of the rearranged fragments. A deletion from the switch mu region (S mu) to the first constant mu exon (C mu 1) and a second deletion from the second constant mu exon (C mu 2) to the gamma 3 region (gamma 3) was detected in the low-grade case. In the high-grade case, there was a deletion of the IgH intronic enhancer (E mu) and a 336-base pair (bp) insertion into the S mu region of a gene (KIAA0307) normally located at 15q24. These data demonstrate for the first time the ability of MALT lymphoma cells to undergo aberrant isotype switch recombinations, which might be directly involved in the development or progression of malignancy.

Isolation and characterisation of recombination events involving immunoglobulin heavy chain switch regions in multiple myeloma using long distance vectorette PCR (LDV-PCR).

Immunoglobulin class switching occurs as a result of recombination between pairs of switch region sequences located 5' to each constant heavy chain gene except Cdelta. In the B cell neoplasm multiple myeloma, tumour cells have generally undergone class switching and often contain oncogenic sequences translocated into switch regions, presumably as a result of aberrant switch recombination. We have developed a method (LDV-PCR) which combines long distance PCR with one-sided vectorette PCR that is capable of detecting and isolating both normal and aberrant switch recombination breakpoints from multiple myeloma cell lines and primary multiple myeloma tumour material. Using LDV-PCR we have directly cloned the translocation breakpoints present in two multiple myeloma cell lines and isolated a normal productive switch recombination event from a primary tumour. Furthermore, we have isolated a novel translocation t(14;22)(q32;q12) from a primary tumour sample and have demonstrated that internal deletions within switch regions can occur in multiple myeloma cells. Compared to a Southern blotting approach, LDV-PCR is simpler and more rapid to perform, allows the simultaneous detection and isolation of recombination events, and can also be applied to amounts of DNA which are too low to permit the conventional cloning of recombination breakpoints.

G4 DNA Binding by LR1 and Its Subunits, Nucleolin and hnRNP D, A Role for G-G pairing in Immunoglobulin Switch Recombination

The immunoglobulin heavy chain switch regions contain multiple runs of guanines on the top (nontemplate) DNA strand. Here we show that LR1, a B cell-specific, duplex DNA binding factor, binds tightly and specifically to synthetic oligonucleotides containing G-G base pairs (KD </= 0.25 nM). LR1 also binds to single-stranded G-rich sequences (KD approximately 10 nM). The two subunits of LR1, nucleolin and hnRNP D, bind with high affinity to G4 DNA (KD = 0.4 and 0.5 nM, respectively). LR1 therefore contains two independent G4 DNA binding domains. We propose that LR1 binds with G-G-paired structures that form during the transcription of the S regions that is prerequisite to recombination in vivo. Interactions of donor and acceptor S regions with subunits of the LR1 could then juxtapose the switch regions for recombination.

Developmental specificity of immunoglobulin heavy chain switch region recombination activities

To understand the regulation of enzymes that carry out immunoglobulin heavy chain class switch recombination, we have assayed recombination of extrachromosomal substrates carrying switch region sequences in cell lines representing different stages of lymphoid cell development. Both pre-B and mature B cell lines supported switch substrate recombination, but B cell lines derived from later stages of cell development did not. Recombination did not occur in an erythroid or a macrophage line. Most recombination junctions in the substrates recovered from transfection of pre-B and B cells mapped to heterogeneous sites within the Sμ and Sγ regions, as do chromosomal switch junctions. Some recombination did occur in T cell lines, but most recombination junctions involved an upstream promoter and did not map preferentially to S regions. Culture of the pre-B cell lines PD31 and 70Z 3 with LPS increased recombination two-fold, to levels approaching those observed in LPS-cultured primary B cells. These results show that the full complement of factors necessary for switch recombination is present only in cells representing a limited spectrum of B cell development and that LPS, which can activate resting splenic B cells to carry out chromosomal recombination, can also stimulate recombination activity in immortalized pre-B cell lines.

Promiscuous translocations into immunoglobulin heavy chain switch regions in multiple myeloma.

In multiple myeloma, karyotopic 14q32 translocations have been identified at a variable frequency (10-60% in different studies). In the majority of cases, the partner chromosome has not been identified (14q+), and in the remaining cases, a diverse array of chromosomal partners has been implicated, with 11q13 being the most common. We developed a comprehensive Southern blot assay to identify and distinguish different kinds of immunoglobulin heavy chain (IgH) switch recombination events. Illegitimate switch recombination fragments (defined as containing sequences from only one switch region) are potential markers of translocation events into IgH switch regions and were identified in 15 of 21 myeloma cell lines, including seven of eight karyotyped lines that have no detectable 14q32 translocation. From all nine lines or tumor samples analyzed further, cloned illegitimate switch recombination fragments were confirmed to be IgH switch translocation breakpoints. In three of these cases, the translocation breakpoint was shown to be present in the primary tumor. These translocation breakpoints involve six chromosomal loci: 4p16.3 (two lines and the one tumor); 6; 8q24.13; 11q13.3 (in three lines); 16q23.1; and 21q22.1. We suggest that translocations into the IgH locus (i) are frequent (karyotypic 14q32 translocations and/or illegitimate switch recombination fragments are present in primary tumor samples and in 19 of 21 lines that we have analyzed); (ii) occur mainly in switch regions; and (iii) involve a diverse but nonrandom array (i.e., frequently 11q13 or 4p16) of chromosomal partners. This appears to be the most frequent genetic abnormality in multiple myeloma.

Promiscuous translocations into immunoglobulin heavy chain switch regions in multiple myeloma

Promiscuous translocations into immunoglobulin heavy chain switch regions in multiple myeloma

Chromosomal translocations cause deregulated BCL6 expression by promoter substitution in B cell lymphoma.

The BCL6 gene codes for a zinc-finger transcription factor and is involved in chromosomal rearrangements in 30-40% of diffuse large-cell lymphoma (DLCL). These rearrangements cluster within the 5' regulatory region of BCL6 spanning its first non-coding exon. To determine the functional consequences of these alterations, we have analyzed the structure of the rearranged BCL6 alleles and their corresponding RNA and protein species in two DLCL biopsies and one tumor cell line which carried the t(3;14)(q27;q32) translocation involving the BCL6 and immunoglobulin heavy-chain (IgH) loci. In all three cases, the breakpoints were mapped within the IgH switch region and the BCL6 first intron, leading to the juxtaposition of part of the IgH locus upstream and in the same transcriptional orientation to the BCL6 coding exons. An analysis of cDNA clones showed that these recombinations generate chimeric IgH-BCL6 transcripts which initiated from IgH germline transcript promoters (I mu or I gamma 3), but retain a normal BCL6 coding domain. In the tumor cell line, the chimeric I gamma 3-BCL6 allele, but not the germline BCL6 gene, was transcriptionally active and produced a normal BCL6 protein. These findings indicate that t(3;14) translocations alter BCL6 expression by promoter substitution and imply that the consequence of these alterations is the deregulated expression of a normal BCL6 protein.

Differences in the molecular structure of c-myc-activating recombinations in murine plasmacytomas and precursor cells.

The translocation of c-myc on chromosome (chr.) 15 to an immunoglobulin heavy-chain switch region on chr. 12 is the critical oncogenic step in pristane-induced plasmacytoma (PCT) development in BALB/cAnPt mice. Applying a recently developed PCR method, we have been able to detect the most commonly occurring illegitimate recombinations between alpha-chain switch region (S alpha) and c-myc in preneoplastic B cells residing in mesenteric oil granuloma (OG) tissues 7-30 days postpristane. In this study, we compare the nucleotide sequences at the S alpha/c-myc breaksites on both the c-myc-activating chr. 12+ and the reciprocal chr. 15- from eight transplanted PCTs, seven primary PCTs, and five OGs that contained six B-cell clones. These junction sequences revealed a remarkable diversity of S alpha/c-myc recombinations. In nine cases--four PCTs and five B-cell clones--nearly precise reciprocal exchanges with a loss of only 3-35 bp in c-myc were found. Large deletions in c-myc that removed 369-878 bp were observed in seven PCTs but not in early B cells. Duplications of c-myc ranging from 103 to 229 bp were also restricted to PCTs and noticed in four cases. Clonally related but different reciprocal recombinations, 38 bp apart on chr. 12+ and 15 bp apart on chr. 15-, were isolated from two different specimens of the same OG tissue from a BALB/c mouse 30 days postpristane. A second OG from another 30-day mouse yielded four recombinational fragments--two clonally related chr. 12(+)-specific fragments and two chr. 15(-)-specific fragments--one of which carried a 143-bp insertion of a microsatellite at the breaksite. We suggest that the initial recombinational break-point regions between S alpha and c-myc in plasmacytoma precursor cells at the time of immunoglobulin heavy-chain switching are intrinsically labile and characterized by a persisting instability of c-myc, which can result in large secondary deletions of c-myc.

The polymorphism of the immunoglobulin heavy chain switch region gene in IgA nephropathy, Henoch-Schönlein nephritis and idiopathic nephrotic syndrome.

It is well known genetic predisposition may play an important role in the pathogenesis of renal diseases. Recently, there has been some controversy about the possible role of the polymorphism of the immunoglobulin heavy chain switch region gene. We have studied this gene by SstI restriction fragment length polymorphisms using DNA from 41 children with IgA nephropathy, 44 with Henoch-Schönlein nephritis and 60 with idiopathic nephrotic syndrome. There was no association of specific genotype with these diseases, in contrast to previous reports. These results are probably due to the differing genetic background of Japanese and Caucasoid patients as far as the switch region is concerned; no switch region genotype constitutes a genetic risk factor for the Japanese in these diseases.