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Mice with transgenes containing an antibody H chain V region (VHDJH) gene were used in an analysis of the cis-acting elements required for hypermutation of immunoglobulin (Ig) V genes. These transgenes can somatically recombine with endogenous IgH DNA, leading to the formation of functional heavy (H) chains partially encoded by the transgenic VHDJH. The transgenomes in the five different lines of mice analyzed contain as little as 150 bp, and as much as 2.8 kb of natural DNA flanking the 5' side of the VH and either 1.5 or 2.3 kb (including the intronic enhancer and 5' matrix attachment region [MAR]) flanking the 3' side of VH. Hybridomas were constructed from immunized transgenic mice, and transgenes present in these hybridomas that had or had not recombined to form functional H chain loci were sequenced. The data obtained show that: (a) the recombined transgenes contain hypermutated VH genes; and (b) among such transgenes, even those containing only 150 bp of natural VH 5' flanking sequence and several kilobases of 5' plasmid vector sequence display a frequency, distribution, and type of mutation characteristic of conventional IgH loci. The data also indicate that transgenic VHDJH genes that have not recombined with endogenous IgH DNA are not substrates for hypermutation, even if they are flanked by 2.8 kb of natural 5' DNA, and 2.3 kb of natural 3' DNA, including the JH2-JH4 region, a MAR, and the intronic enhancer. Collectively, the data suggest that sequences 5' of the VH promoter are dispensable, a VH promoter and the intronic IgH enhancer region are not sufficient, and a region(s) within or 3' of the IgH constant region locus is requisite, for hypermutation of Ig VH transgenes.

the journal of experimental medicine/the journal of experimental medicine

Hypermutation is observed only in antibody H chain V region transgenes that have recombined with endogenous immunoglobulin H DNA: implications for the location of cis-acting elements required for somatic mutation.