Expression of 12 rabbit IgA C alpha genes as chimeric rabbit-mouse IgA antibodies.
Serologic analysis of rabbit secretory IgA initially identified two subclasses of IgA, IgA-f and IgA-g. Recent molecular genetic studies have resulted in the identification and cloning of 13 genes encoding the constant region (C) of rabbit IgA heavy chains. Each of these 13 C alpha genes, C alpha 1-C alpha 13, was subcloned into an expression vector containing the VDJ (V, variable; D, diversity; J, joining) gene of a dansyl (DNS)-binding hybridoma antibody. The alpha heavy-chain constructs were transfected into SP2/0 cells producing murine light chains with specificity for DNS. Of the 13 resulting transfectomas, 12 were shown by ELISA to secrete DNS-binding chimeric rabbit-mouse IgA molecules. By immunoblot analysis, the 12 IgA-producing transfectomas were shown to secrete alpha chains ranging in size from 60 to 72 kDa. These data suggest that rabbit IgA may be composed of as many as 12 IgA isotypes. This is in marked contrast to mouse and human, in which only 1 and 2 IgA isotypes, respectively, are found. Serologic analyses, using anti-IgA-f and anti-IgA-g alloantisera, revealed that 11 of the 12 transfectoma IgAs reacted with anti-IgA-f and not with anti-IgA-g antibodies and that one reacted with anti-IgA-g and not with anti-IgA-f antibodies. Each of the IgA-producing transfectomas was cocultured with a Madin-Darby canine kidney cell line expressing the rabbit polymeric immunoglobulin receptor, and the transcytosed IgA antibodies were analyzed by immunoblots to determine whether they associated with secretory component (SC) through covalent or noncovalent interactions. Each of the 11 IgA-f isotypes was shown to bind SC by a disulfide linkage, whereas the single IgA-g isotype appeared to bind SC through noncovalent interactions only.
Schneiderman
R D
RD
Department of Microbiology and Immunology, University of Illinois, College of Medicine, Chicago 60680.
Hanly
W C
WC
Knight
K L
KL
D000818:Animals; D002460:Cell Line; D002678:Chimera; D004797:Enzyme-Linked Immunosorbent Assay; D005803:Genes, Immunoglobulin; D005822:Genetic Vectors; D006239:Haplotypes; D015151:Immunoblotting; D007070:Immunoglobulin A / Q000032:analysis / Q000145:classification / Q000235:genetics*; D007127:Immunoglobulin Constant Regions / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D051379:Mice; D010954:Plasmacytoma; D011817:Rabbits; D014162:Transfection
D007070:Immunoglobulin A; D007127:Immunoglobulin Constant Regions; D007143:Immunoglobulin Heavy Chains