The Major Histocompatibility Complex (MHC) in the cat (FLA) contains highly polymorphic genes, many of which are critical in regulating the immune response. The cat MHC contains up to three DRA and DRB genes, but lacks DQ and DP genes. We have investigated FLA-DRB alleles in a variety of domestic and wild felids.

DNA samples from 109 domestic cats and 67 wild cats (from 16 different species) were characterised using DNA sequencing and cloning, and Reference Strand-mediated conformation analysis (RSCA).

Table 1: Samples Available for this Study

Method

Reference strand-mediated conformation analysis:

 Used to analyse variations in the DNA conformation of different alleles using capillary electrophoresis

 Uses a fluorescently labelled "reference" (FLR), generated by PCR

 FLR is hybridised with the PCR products from test samples:-> duplexes form.

 Duplexes are run through capillaries past a laser (AB! 3100 capillary analyser)

 Detected duplexes have the FLR sense strand in common, but the molecular conformation of each duplex is unique, because each antisense strand is different

 Therefore duplex mobility is unique:

 -> this gives separation of individual alleles, see Figures 1 & 2.

Results

 RSCA is a good way of characterising FLA-DRB alleles in all cats

 Cats can have 1-6 alleles, confirming there are up to three FLA-DRB genes

 Family studies indicate that there are different numbers of genes on different haplotypes, see Figure 3

 33 different alleles found in domestic cats to date

 Another 25 in 15 different felid species to date

 All the sequences found to date fit in the same allelic series

 There are no species specific DNA motifs

 Phylogenetic analysis shows the alleles do not fall into groups by species, see Figure 4

Figure 1. Typical RSCA Results for Two Domestic Cats

Figure 2. Typical RSCA Results for Some Big Cats

Results

These data indicate that individual cats can have one to six alleles present, suggesting that different haplotypes may carry one, two or three DRB genes.

To date, we have been unable to assign alleles to specific loci. Fifty of our samples are from family members, but it has not proved easy to assign haplotypes. This may be complicated by the possibility that available primers do not amplify all alleles.

Interestingly there was some sharing of alleles between different species. One allele was found in both lions and jaguars, and another in both Amur and Persian leopards. The Indian desert cat shared an allele with the domestic cat.

Take Home Messages

 FLA-DRB in the cat can be characterised by RSCA

 There is lots of diversity!

 All the DRB alleles found in all cats fit in the same allelic series

 Different haplotypes carry different numbers of FLA-DRB genes

 Further work is needed to dissect the complexity of the FLA-DRB genes