Reactivation by exon shuffling of a conserved HLA-DR3-like pseudogene segment in a New World primate species

Abstract

The common marmoset (Callithrix jacchus), a New World monkey species with a limited MHC class II repertoire, is highly susceptible to certain bacterial infections. Genomic analysis of exon 2 sequences documented the existence of only one DRB region configuration harboring three loci. Two of these loci display moderate levels of allelic polymorphism, whereas the -DRB*W12 gene appears to be monomorphic. This study shows that only the Caja-DRB*W16 and -DRB*W12 loci produce functional transcripts. The Caja-DRB1*03 locus is occupied by a pseudogene, given that most of the transcripts, if detected at all, show imperfections and are present at low levels. Moreover, two hybrid transcripts were identified that feature the evolutionarily conserved peptide-binding motif characteristic for the Caja-DRB1*03 gene. Thus, the severely reduced MHC class II repertoire in common marmosets has been expanded by reactivation of a pseudogene segment as a result of exon shuffling.

Fig. 1.

Caja-DRB alleles detected in a panel of 15 common marmosets. Shading is used to indicate whether the respective allele is detected only on the genomic (exon 2) level (black shading), or on both the genomic and cDNA levels (gray shading). Animals m02040 and m02041 were tested only on cDNA. Because it was impossible to develop -DRB lineage-specific primers that did not crossreact with other DRB cDNAs, real-time PCR results were not reliable. Therefore, transcription levels were determined by counting the number of clones. For every individual, at least 60 clones were analyzed. All transcripts of DRB*W16 and of the DRB1*0307/DRB1*0312 alleles were detected in at least 50% of the clones, and DRB*W12 alleles were detected in ≈25% of the clones. The transcripts of DRB1*03L alleles, however, were found at a much lower extent, namely in <5% of the clones tested.

Fig. 2.

Alignment of deduced Caja-DRB amino acid sequences, given in conventional one-letter code. A dash or dot indicates identity with the consensus or deletion of an amino acid, respectively, whereas ∗ indicates that this part of the sequence is unknown. Amino acids or deletions, which are typical for the -DRB1*03 and -DRB*W16 lineages, are shown in yellow or blue, respectively. Consequently, the names of the hybrid genes, -DRB1*0307 and -DRB1*0312, are shown in both blue and yellow. L, leader peptide and first four amino acids of the coding sequence; β₁, β₁ domain; β₂, β₂ domain; TM, transmembrane region; CYT, cytoplasmic tail; 3′UT, 3′ untranslated region.

Fig. 3.

Phylogenetic tree constructed according to the neighbor-joining method, illustrating the evolutionary relationships between alleles of various primate species. Bootstrap values are indicated. The alleles depicted in this tree were selected from this study or from the ImMunoGeneTics/Non-Human Primates database (23). Dog leukocyte antigen (DLA) -DRB is chosen as outgroup. Caja, Callithrix jacchus; Saoe, Saguinus oedipus; Gogo, Gorilla gorilla; Mamu, Macaca mulatta; Mane, Macaca nemestrina.

Fig. 4.

Schematic representation of the exon-shuffling event producing a novel Caja-DRB gene.