Somatic hypermutation of immunoglobulin ĸ light chain genes

Goyenechea Corzo, Beatriz (1996). Somatic hypermutation of immunoglobulin ĸ light chain genes. PhD thesis The Open University.



Somatic hypermutation of immunoglobulin genes is a tightly controlled, site directed process. The nucleotide changes are targeted in the region surrounding the rearranged V gene segment of the heavy and light chain loci. The targeting of somatic mutation does neither require V region including the Vĸ promoter specific features nor a specific distance between the C ĸ and the 3' enhancer. On the other hand, removal of either the ĸ 3' enhancer or the ĸ intron enhancer/M AR causes a drastic reduction in hypermutation, which is particularly marked in the latter case. In the work described in this dissertation I have used transgenic mice carrying immunoglobulin ĸ transgenes with deleted regulatory elements, to make a tighter delineation of which DNA regions within the ĸ locus are important for hypermutation. In addition I have used transgenic mice to study the role of the primary sequence and the DNA secondary structure in this process.

This thesis describes the generation of artificial Ig genes constructs to make transgenic lines. ĸ transgenes were characterized by Southern blot and by ELISA in the seven mouse lines obtained. The role of the 3' enhancer core in the transcription of the ĸ chain was also studied in stable transfected S I07 mouse myelom a cells.

The analysis of the transgenes carrying deletions of regulatory elements revealed that hypermutation was impaired (but not severely affected) when the intron enhancer was deleted. This also was the case when the flanking regions of the 3' enhancer core were deleted. Hypermutation was severely affected (but not abolished) when the core of the 3' enhancer was removed. This reflects the relative roles of both components of the 3' enhancer region in gene expression. The matrix attachment region seems to be essential, since its deletion abolished hypermutation, but not expression, since abundant ĸ mRNA was still produced. Furthermore these studies showed that the 3' enhancer can drive hypermutation from a position upstream of the Vĸ region.

The major role of the primary sequence in the formation of hotspots and the mutational machinery preferences for AGY encoded Ser and the bias against TCA encoded Ser as target for hypermutation were disclosed with the analysis of the transgenes carrying VĸOx1 gene with modifications in the CDR1 sequence. However, whilst hairpin loops do not define which base is going to be targeted, they seem to have a role in the accessibility of the 'favoured' nucleotides for hypermutation.

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