Deregulation of the proto-oncogene c-myc through t(8;22) translocation in Burkitt's lymphoma.

In Burkitt's lymphoma (BL) cells the proto-oncogene c-myc is juxtaposed to one of the immunoglobulin (Ig) loci on chromosomes 2, 14, or 22. The c-myc gene becomes transcriptionally activated as a consequence of the chromosomal translocation and shows preferential usage of promoter P1 over P2, a phenomenon referred to as promoter shift. In order to define the responsible regulatory elements within the Ig lambda locus, we studied the effect of the human Ig lambda enhancer (HuE lambda) on c-myc expression after stable transfection into BL cells. A 12 kb genomic fragment encompassing HuE lambda, but not HuE lambda alone, strongly activated c-myc expression and induced the promoter shift. To identify additional elements involved in c-myc deregulation, we mapped DNaseI hypersensitive sites within the 12 kb lambda fragment on the construct. Besides one hypersensitive site corresponding to HuE lambda, three additional sites were detected. Two of these elements displayed enhancer activity after transient transfection. The third element did not activate c-myc transcription, but was required for full c-myc activation and promoter shift. Deletion analyses of the c-myc promoter identified the immediate promoter region as sufficient for activation by the Ig lambda. locus, but also revealed that induction of the promoter shift requires additional upstream elements.

Tissue-specific deoxyribonuclease I-hypersensitive sites in the vicinity of the immunoglobulin C lambda cluster of man.

During B cell development, the onset of DNA rearrangements, expression, and somatic hypermutation of Ig genes are regulated through the complex interaction of cis-acting elements with trans-acting factors. Our aim is to identify DNA elements required during activation of the human Ig lambda light chain genes. Determination of deoxyribonuclease (DNase) I-hypersensitive sites in complex regulated genes can lead to the identification of sequence elements which would have been overlooked by employing transient transfection protocols. We have therefore investigated the chromatin structure of human J-C lambda genes and identified three DNase I-hypersensitive sites (HSS-1, -2, and -3) within an 8-kb region downstream of the J-C lambda 7 gene. HSS-2 and HSS-3 are B cell specific. The DNase I-hypersensitive sites are also present in kappa-producing cell lines which have not rearranged the Ig lambda locus and produce germ-line J-C lambda transcripts. We conclude that in mature B cells, both kappa and lambda loci are in an active structure regardless of the type of light chain they produce. This suggests that the chromatin structure of both loci is opened early in B cell development and that the active structure persists in mature B cells. The observed temporal order (first kappa, then lambda) of activation can be explained by consecutive synthesis of the appropriate regulating factors and the tight regulation of the recombination machinery through the products of L chain gene rearrangements.

The immunoglobulin lambda light chain enhancer consists of three modules which synergize in activation of transcription.

V(D)J rearrangement, high level expression and somatic hypermutation of assembled Ig genes is tightly controlled by a number of regulatory sequence elements located in the vicinity of the J-, (D)-, and C-gene segments. During B cell maturation these elements become accessible to binding of trans-acting factors, reflecting the opening of the chromatin structure through an as yet unidentified mechanism. The mapping of regions of an altered chromatin structure (DNase I hypersensitivity) therefore is a powerful approach in identifying regulatory sequence elements. We here show that the human Ig lambda enhancer consists of three modules previously identified by us as DNase I-hypersensitive sites HSS-1, -2, and -3. The three sequence elements synergize in transcriptional activation of a reporter gene and together constitute a powerful tissue-specific enhancer which is a much stronger transcriptional activator than the kappa enhancers alone or in combination. We further show that the accessibility of the kappa and lambda enhancer elements for DNase I in the chromatin of a pre-B cell line (207) correlates with the transcriptional enhancer activities of kappa and lambda enhancer constructs. This finding is in support of an ordered model for Ig light chain activation (kappa before lambda).