Isolation of carp genes encoding major histocompatibility complex antigens

In the evolution of the adaptive immune system unique to vertebrates, teleost fish occupy the critical position. This is the most primitive class of lower vertebrates in which the capacity for acute allograft rejections can be demonstrated, thus suggesting the presence of major histocompatibility complex (MHC) antigens and, therefore, T cells. Here, we report the identification of two putative MHC-antigen-encoding sequences in the carp Cyprinus carpio. One, identified as TLAI alpha-1, had reasonable homology to MHC class I heavy chains of mammalian and avian species, while the other, identified as TLAII beta-1, was homologous to MHC class II beta chain of the aforementioned higher vertebrates. For these isolations of fish MHC genes, we have identified two highly conserved amino acid sequence blocks surrounding two cysteine residues in the second domain of MHC class II beta chains as well as the third domain of class I heavy chains of humans, mice, and chickens. Two kinds of mixed oligonucleotide probes corresponding to these two regions were synthesized. The carp genomic DNA was subjected to amplification by polymerase chain reaction using the above two synthetic DNA fragments as primers. Subsequently, two different DNA sequences sandwiched by these primers were isolated from the amplified products. Their use as secondary probes led to the identification of TLAI alpha-1 and TLAII beta-1. We also discuss the applicability of the above approach for isolation from lower vertebrates of other genes belonging to the immunoglobulin super-family as well as the evolutionary origin of vertebrate MHC antigens.