Structural and genetic studies of chicken 7S immunoglobulin allotypes

Wakeland, Edward K.
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Six chicken 7S immunoglobulin (Ig) allotypic specificities were demonstrated with antisera produced in highly inbred chickens. A radioimmunoassay which utilized intact, radiolabeled 7S Ig alloantigens was developed, and was used for the structural and genetic analysis of these specificities. The binding of dinitrophenylated alloantiserum (DAA) with radiolabeled alloantigen was demonstrated by the addition of rabbit anti-dinitrophenyl antiserum followed by precipitation of complexes with sheep anti-rabbit γ-globulin antiserum. Binding inhibition assays with the DAA method detected allotypes on 2 to 5 ng of 7S Ig alloantigen. Structural analyses of the six allotypic specificities were performed by direct binding and binding inhibition analysis with the DAA assay. Specificities CS-1.1 and CS-1.2 were present on papain-produced Fab fragments and isolated 7S Ig heavy (H) chains; but not on light (L) chains, Fc fragments, or 17S Ig. For both specificities H chains reassociated with L chains were three- to five-fold better inhibitors on a weight basis than isolated H chains. Regression coefficients calculated from binding inhibition curves indicated that the avidity of H chains for specific alloantisera was significantly increased following reassociation with L chains, suggesting that the conformation of CS-1.1 and CS-1.2 determinants may depend in part upon Hand L chain interactions. Specificities CS-1.3 and CS-1.4 were detected on isolated 7S Ig H chains; but not on L chains, Fab, Fc, or l7S Ig. In contrast to the results with CS-1.1 and CS-1.2, reassociation of Hand L chains did not significantly increase the inhibitory activity or binding avidity of CS-1.3 and CS-1.4 for specific alloantisera. Specificity CS-1.5 was also present on intact 7S Ig, but absent from Fab, Fc, or 178 Ig. A nonhomologous cross reaction was observed in binding inhibition assays with UCD 2 anti-CS-1.2 alloantiserum. The cross-reacting specificity, designated CS-1.6, was found on 7S Ig and was absent from l7S Ig enriched preparations. These data indicate that at least two regions of the chicken 7S Ig H chain contain genetic variations: (1) specificities CS-1.1 and CS-1.2 are located in the Fd portion of the H chain, perhaps in a region analogous to the CH1 domain of mammalian IgG; and (2) specificities CS-1.3, CS-1.4, and CS-1.5 are located in a region of the H chain which is sensitive to papain digestion. Among F2 progeny, these specificities segregated in a manner statistically indistinguishable from that expected of codominant alleles of a "Mendelian" gene at an autosomal locus. This gene, designated CS-1, occupied a locus which was not closely linked to any of five chicken blood group loci. Combinations of 7S Ig allotypic specificities were inherited as stable phenogroups among the F2 progeny of inbred chicken lines, and direct binding analysis with the DAA assay indicated that the specificities forming phenogroups were present on the same 7S Ig molecules. These results also demonstrated that more than 94% of the serum 7S Ig contained the products of the CS-1 gene. In FI hybrids with the genotype CS-1.1,1.3/1.2, two populations of serum 7S Ig molecules were detected by direct and sequential binding analysis with alloantisera. One population of 7S Ig contained specificities CS-1.1 and CS-1.3, but not CS-1.2; while the second population carried only specificity CS-1.2. Therefore, each population was exclusively the product of one parental allele. Consistent with a genetic regulatory mechanism involving allelic exclusion, no 7S Ig containing allotypes produced by both alleles was detected. A survey of 43 inbred lines derived from four sources in the United States for 7S Ig allotypes revealed considerable genetic polymorphism. A minimum of ten alleles of the CS-1 gene were found as unique combinations of CS-1 specificities. A system of nomenclature for CS-1 alleles was developed and six inbred lines were designated as prototype lines. The remaining four CS-1 alleles occurred only in inbred lines which were polymorphic for 7S Ig allotypic specificities.
Thesis (Ph. D.)--University of Hawaii at Manoa, 1976.
Bibliography: leaves 141-150.
xv, 150 leaves ill
Immunoglobulins, Immunogenetics
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Theses for the degree of Doctor of Philosophy (University of Hawaii at Manoa). Microbiology; no. 953
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