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Allelic Variation at the V_Ha Locus in Natural Populations of Rabbit (Oryctolagus cuniculus, L.)¹

Pedro J. Esteves^*,, Dennis Lanning, Nuno Ferrand^*, Katherine L. Knight, Shi-Kang Zhai and Wessel van der Loo^2,*,

^* CIBIO, Centro de Investigação em Biodiversidade e Recursos Genéticos, Campus Agrário de Vairão, Rua Padre Armando Quintas, Vairão, Portugal, and Departamento de Zoologia e Antropologia, Faculdade de Ciências da Universidade do Porto, Porto, Portugal; Institute of Molecular Biology and Biotechnology, Vrije Universiteit Brussel, Brussels, Belgium; and Department of Microbiology and Immunology, Loyola University Chicago, Maywood, IL 60153

The large interallelic distances between the three rabbit Ig V_Ha lineages, a1, a2 and a3, suggest that the persistence time of the V_Ha polymorphism could amount to 50 million years, which is much longer than that of MHC polymorphisms. Rabbit originated in the Iberian Peninsula where two subspecies coexist, one of which is confined to Southwestern Iberia (Oryctolagus cuniculus algirus). We studied the V_H loci in the original species range to obtain a better understanding of the evolutionary history of this unusual polymorphism. Serological surveys revealed that sera from the subspecies algirus, when tested with V_Ha locus-specific alloantisera, showed either cross-reactivity ("a-positive" variants) or no reaction at all ("a-blank"). Using RT-PCR, we determined 120 sequences of rearranged V_H genes expressed in seven algirus rabbits that were typed as either a-positive or a-blank. The data show that the V_H genes transcribed in a-positive rabbits are closely related to the V_H1 alleles of domestic rabbits. In contrast, a-blank rabbits were found to preferentially use V_H genes that, although clearly related to the known V_Ha genes, define a new major allotypic lineage, designated a4. The a4 sequences have hallmark rabbit V_Ha residues together with a number of unprecedented amino acid changes in framework region 2 and 3. The net protein distances between the V_Ha4 and the V_Ha1, a2, and a3 lineages were 20, 29, and 21% respectively. We conclude that at least four distantly related lineages of the rabbit V_Ha locus exist, one of which seems to be endemic in the Iberian range.