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SPOTLIGHT 18 of the first Draft of the Human Genome, Identification of the the Nobel Prize for Medicine in 1987

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SPOTLIGHT 18 of the first Draft of the Human Genome, Identification of the the Nobel Prize for Medicine in 1987 Leukemia (2002) 16, 973–984 2002 Nature Publishing Group All rights reserved 0887-6924/02 $25.00 www.nature.com/leu REVIEW The configuration of the immunoglobulin genes in B cell chronic lymphocytic leukemia MJS Dyer1 and DG Oscier2 1Department of Haematology, University of Leicester, Leicester, UK; and 2Department of Haematology, Royal Bournemouth Hospital, Bournemouth, UK B cell chronic lymphocytic leukemia (CLL) lacks a consistent associated with loss of expression of some genes, but possibly genetic abnormality. However, immunoglobulin VH gene seg- over-expression of others.8 ment mutation analysis has provided insights into the patho- genesis of these diseases and allowed the development of Given these difficulties, an alternative approach has been powerful prognostic markers. Immunoglobulin gene to search for abnormalities within genes of importance in the chromosomal translocations are rare in CLL and involve a dis- pathogenesis of other subtypes of B cell malignancy. The B tinct subset of genes including BCL3, BCL11A and CCND2. cell non-Hodgkin’s lymphomas (B-NHL) are in many BCL2 translocations in CLL appear to arise via a different instances, characterized by specific and recurrent chromo- mechanism from comparable translocations seen in B cell non- somal translocations involving the immunoglobulin (IG) Hodgkin lymphoma. 9–11 Leukemia (2002) 16, 973–984. DOI: 10.1038/sj/leu/2402528 loci. Molecular cloning of such translocations has allowed, Keywords: chronic lymphocytic leukemia; immunoglobulin genes; and continues to allow, cloning of genes intimately involved VH mutations; BCL6 mutations; chromosomal translocations in the development of both normal and malignant B cells.12–15 These genes may be involved in the pathogenesis of B cell malignancies, not only through chromosomal translocation, Introduction but also through their direct involvement in genomic amplifi- cations and through changes in methylation. Moreover, Despite much work, the molecular pathogenesis of B cell molecular analysis of the breakpoint sequences allows identi- chronic lymphocytic leukemia (CLL) remains obscure. The fication of the mechanisms and timing of chromosomal trans- 10,16 classical cytogenetic-lead approach to the molecular dissec- locations in B cell development. tion of malignancy has thus far failed to allow the identifi- Here, we review data concerning the configuration of the cation of the major ‘players’. The reasons for this reflect not IG genes in CLL and how studies of these genes have shed only the difficulties in obtaining metaphases from patients light on the pathogenesis and progression of CLL. with CLL, but also the lack of a consistent and a genetically amenable lesion. Cytogenetic analysis of CLL supplemented with comparative genomic hybridization and interphase FISH Somatic recombination and hypermutation of the IG genes studies, have shown recurrent deletions involving regions in normal B cells such as 6q21, 11q22-q23, 13q14.3 and 17p13.1.1 According to the Knudson hypothesis, such deletions may mark the pres- The origins of antibody diversity reflect the actions of three ence of tumor suppressor genes (TSGs) with deletion of one fascinating genetic mechanisms summarized with reference to allele and mutation of the other. Whilst this hypothesis has the immunoglobulin heavy chain (IGH) locus at the very telo- worked well in the isolation of inherited cancer genes, the mere of the long arm of chromosome 14 at chromosome number of somatically mutated TSGs remains small. There are 14q32.33 in Figure 1 and reviewed in Refs 10 and 17 and a variety of possible reasons for this.2 Firstly, somatically Refs therein. acquired deletions are frequently large, spanning several Firstly, somatic recombination of the dispersed IG gene seg- megabases of DNA and may be secondary events, present ments is required to produce a functional IG gene. This was only in a fraction of malignant cells. Even with the completion first demonstrated by Susumu Tonegawa, for which he won SPOTLIGHT 18 of the first draft of the human genome, identification of the the Nobel prize for Medicine in 1987. During the early key gene within the involved region (such as the commonly stages of B cell differentiation within the bone marrow, diver- deleted region of 13q14 in CLL) may therefore be extremely sity (D) and then variable (V) region gene segments undergo difficult.3–5 Alternatively, for some genes, loss of one copy DNA sequence specific recombination with the joining (J) without mutation of the remaining allele (haploinsufficiency) region segments in order to produce a contiguous VDJ gene. may be sufficient, whilst for other TSGs, epigenetic factors, This is mediated by a complex of enzymes, including RAG1 such as promoter methylation may also contribute to loss of and RAG2, which bind to and cleave specific DNA expression.6,7 Furthermore, in CLL, some 13q14 deletions may sequences. Part of the genetic diversity arises through the dif- in fact be associated with chromosomal translocations, sug- ferent junctions arising from the assortment of the various V, D gesting that some of these events are complex and may be and J segments. Cell surface expression of functional antibody molecules is necessary for the continued survival of early pre- B cells. Failure to undergo recombination successfully due to out-of-frame recombination, results in cell death by apoptosis. Correspondence: MJS Dyer, Department of Haematology, University The dependence of B cell survival on maintained surface Ig of Leicester, Robert Kilpatrick Clinical Sciences Building, Leicester 19 Royal Infirmary, PO Box 65, Leicester, LE2 7LX, UK; Fax: 44-116- expression persists until differentiation to memory B cells. 252-3274 After successful recombination, expression of the Received 27 November 2001; accepted 19 February 2002 rearranged VDJH sequences is initially driven by the proximity Configuration of immunoglobulin genes in B cell CLL MJS Dyer and DG Oscier 974 SPOTLIGHT Figure 1 Schema showing the various configurations of the immunoglobulin heavy chain (IGH) locus on chromosome 14 during B cell differentiation. The IGH locus is located at the very telomere of the long arm of chromosome 14. It comprises 52 dispersed variable (VH), 27 diversity (DH) and six joining region (JH) segments. VDJH recombination occurs within the bone marrow and results in the formation of a contiguous gene segment. SHM and CSR occur in the germinal center within the lymph node via a related mechanism. Both involve double- stranded DNA breaks. Switch regions are shown in gold, constant regions in dark blue and enhancer regions in purple. Enhancer regions are ␮ ′ ␣ ␣ ␦ ␥ located in the JH-C intron and 3 of both C 1 and C 2; recently an additional enhancer region has been identified between C and C 3. Transcriptional orientation is shown by horizontal arrows. of the VH promoter to a transcriptional enhancer located some regions are hot spots for SHM are incompletely under- ␮ 28,29 within the intron between the JH segments and C , immedi- stood. ately upstream of the repetitive S␮ region. The anatomy of this In normal B cells, replacement mutations are clustered enhancer, in terms of definition of protein binding sites, has within the CDRs that encode the antibody-combining site, been extensively studied. Apart from its profound effects in rather than the intervening framework regions (FR), which are directing B cell-specific transcription, this small region of responsible for the structural integrity of the variable region. DNA has a number of other interesting properties. It also The ratio of replacement mutations (R) resulting in a change appears to act as an attachment site to the nuclear matrix and in amino-acid content to silent mutations (S), which do not is, as well, an origin of DNA replication.20–22 The intronic affect the protein structure, is therefore higher in CDR than in enhancer of both IG heavy and light chains is necessary for FW regions. The consequence of SHM is the selection of B somatic hypermutation to occur.23 Properties of the enhancer cells that produce high affinity antibodies. Mutated cells that may also account for the differential subnuclear localization produce low affinity antibody or fail to produce antibody, like of the two IGH alleles observed in normal B cells and may the equivalent B cell precursors in the bone marrow, undergo account in part for the allelic exclusion of immunoglobulin apoptosis within the germinal center. The finding of intra- proteins.24 Interestingly, another IG enhancer or possibly an clonal heterogeneity in normal germinal centre B cells indi- IGH locus control region (LCR) has been recently identified cates that these cells may undergo more than one cycle of in a highly unstable region of genomic DNA between the C␦ SHM. and the first downstream C␥ gene, C␥3. This appears to func- Finally, shortly after SHM occurs within the germinal tion in the control of early B cell development although its center,30 the third recombination process, involving transpo- 25 role in mature B cell malignancies has not been investigated. sition of the completed VDJ gene to downstream constant (CH) This region may also contain a number of nuclear matrix region segments is initiated resulting in antibody molecules attachment regions. Breaks in this region are seen in at least with different effector functions. This is known as class switch 10% of cases of B cell precursor ALL, but comparable abnor- recombination (CSR). CSR occurs between highly repetitive malities in CLL have not been detected.26 switch regions, which lie upstream of each of the IG constant Secondly, somatic hypermutation (SHM) is necessary to region genes with the exception of C␦. However, despite the produce antibodies with high affinity for antigen. This occurs highly repetitive nature of individual switch regions, CSR does primarily, but not exclusively, at the complementarity not represent homologous recombination. The discovery that determining regions (CDRs), the antigen-binding sites of the an enzyme with homology to RNA editing enzymes human VH genes.
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