TY - JOUR
T1 - Restriction fragment length polymorphisms and single germline coding region sequence in VH 18 2, a duplicated gene encoding autoantibody
AU - Rubinstein, Daniel B.
AU - Symann, Michel
AU - Stewart, A. Keith
AU - Guillaume, Thierry
N1 - Funding Information:
The human immunoglobulin heavy chain locus has been estimated to be about 2500-3000 kilobases in length and situated at the telomeric end of the long arm of chromosome 14 (Berman et al., 1988; Matsuda et al., 1988). Seven families of VH genes have been identified based on nucleotide homology of greater than 80% between family members (Kabat et a!., 1987; Lee et al., 1987; Berman et al., 1988; Humphries et al., 1988; Mortari et al., 1991). The size of the families, as determined by the complexity seen by hybridization of family probes to genomic DNA, varies from very small (one for VH6) (Berman et al., 1988), to very large (more than 30 for VH3) (Souroujon et al., 1989). Mapping pulsed field el~trophoresis and clones has the VH6 to be the 3’ of the locus (Berman al., 1988) while members of the other VH gene families appear to be highly interspersed. (Kodaira et al., 1986; Berman et al., 1988; Buluwela et al., 1988; Walter et al., 1990, 1991). Polymorphisms are difficult to detect in heterozygous humans because of the complex patterns obtained on *This work was supported by Grants AI26450 and AI28899 from the National Institutes of Health, and by the NATO Exchange Program, and the Association Sportive Contre le Cancer, Brussels. $Author to whom correspondence should be addressed.
PY - 1993/3
Y1 - 1993/3
N2 - In this study, we analyze the restriction fragment length polymorphisms of VH 18 2, a gene encoding the heavy chain of an anti-DNA antibody, and correlate that with the germline sequence of the gene's coding region. Oligonucleotides probes complementary to the CDR1 and to both the 5' and 3' halves of the CDR2 gene segments were hybridized under stringent conditions to genomic DNA digested with Xbal, and yielded polymorphic bands of 26, 24 and 18 kb with all three probes. Individuals had either one, two, or three bands in common in their genomic DNA, indicating duplication of VH 18 2 genes at two sites within the IgH locus. While it is difficult to say which of the three polymorphic gene segments constitute an allelic pair, the 26 and 24 kb fragments were the most commonly seen (97% of individuals had one or both). VH 18 2 is known to be over-represented in the expressed repertoire with very little nucleotide divergence from the germline sequence. In order to determine whether the observed RFLPs are due to sequence polymorphism of the VH 18 2 coding region, and whether differences in the expressed genes arise from somatic mutation, size-selected genomic DNA containing the gene was cloned and sequenced. A single coding region sequence was found in the germline. The results of this study suggest that overexpression of VH 18 2 may in part be due to its duplication. Like other genes encoding autoantibody, which are well conserved, nonpolymorphic and expressed early in programmed immunologic development, VH 18 2 may be instrumental in the establishment or regulation of the immune repertoire.
AB - In this study, we analyze the restriction fragment length polymorphisms of VH 18 2, a gene encoding the heavy chain of an anti-DNA antibody, and correlate that with the germline sequence of the gene's coding region. Oligonucleotides probes complementary to the CDR1 and to both the 5' and 3' halves of the CDR2 gene segments were hybridized under stringent conditions to genomic DNA digested with Xbal, and yielded polymorphic bands of 26, 24 and 18 kb with all three probes. Individuals had either one, two, or three bands in common in their genomic DNA, indicating duplication of VH 18 2 genes at two sites within the IgH locus. While it is difficult to say which of the three polymorphic gene segments constitute an allelic pair, the 26 and 24 kb fragments were the most commonly seen (97% of individuals had one or both). VH 18 2 is known to be over-represented in the expressed repertoire with very little nucleotide divergence from the germline sequence. In order to determine whether the observed RFLPs are due to sequence polymorphism of the VH 18 2 coding region, and whether differences in the expressed genes arise from somatic mutation, size-selected genomic DNA containing the gene was cloned and sequenced. A single coding region sequence was found in the germline. The results of this study suggest that overexpression of VH 18 2 may in part be due to its duplication. Like other genes encoding autoantibody, which are well conserved, nonpolymorphic and expressed early in programmed immunologic development, VH 18 2 may be instrumental in the establishment or regulation of the immune repertoire.
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U2 - 10.1016/0161-5890(93)90070-R
DO - 10.1016/0161-5890(93)90070-R
M3 - Article
C2 - 8096063
AN - SCOPUS:0027465645
SN - 0161-5890
VL - 30
SP - 403
EP - 412
JO - Immunochemistry
JF - Immunochemistry
IS - 4
ER -