RABBIT ALLOTYPES--STRUCTURE, ORGANIZATION AND REGULATED EXPRESSION OF IG GENES

采用分子技术对兔免疫系统基因进行了研究

We studied genes of the rabbit immune system by techniques of molecular biology and immunology. We used anti-RAG-1 and 2 antibodies in studies of developing rabbit appendix tissues. The surface markers CD43 and IgM distinguished appendix cell populations from 6 to 9-week-old rabbits that contained RAG-1 and RAG-2 proteins. The appearance of CD43 during different stages of B-cell maturation may be related to the function of the appendix as a site of both B-cell development and diversification. In the ali mutant rabbit, a small deletion at the 3' end of the V/H gene cluster led to loss of V/H1 and one V/H pseudogene. Although homozygous mutant ali/ali rabbits lack the V/H1a2 gene, B cells with a2-like surface Ig develop and expand in numbers. We sequenced immunoglobulin heavy chain variable regions (V/H) from expressed mRNA of sorted a2-positive and negative appendix B cell populations from young ali rabbits. The a2 positive cells appear to have rearranged V/H4, the first functional gene in the mutants' V/H cluster. Sequence alterations in FR1 and FR3 can be accounted for by gene-conversion-like changes that utilized candidate donor sequences upstream V/H4. Our studies of the appearance of cells bearing a2-like epitopes in the appendix of mutant rabbits suggested that there was positive selection and expansion based on framework region structures (V/Ha allotypes). We suggested that the preferential expansion and survival of B cells based on FR1 and FR3 expression may involve "superantigen"-like interactions with endogenous and exogenous ligands. One endogenous ligand appears to be CD5. In man and mouse, the 3'-most D/H gene, DQ52, is preferentially rearranged early in B-cell development. To test whether this preference for rearranging a D/H gene segment based on 3' end proximity exists in rabbit, we cloned and sequenced the rabbit DQ52 gene. The coding region sequence is identical to a mouse DQ52 but the 3' recombination signal sequence has an atypical nonamer. The DQ52 gene was utilized very infrequently; one VDJ sequence from 28 day fetal liver had 8 bp that matched the germline DQ52 sequence. In contrast to man and mouse, instead of rearranging DQ52 early in B cell ontogeny, rabbits preferentially express another D/H gene (Df) located in the middle of the D/H region about 32 kb upstream of the JH genes. This may correlate with more frequent initial rearrangement of V/H to D/H in rabbit B cells.