THE NATURE OF POLYMORPHISM OF THE HLA-DRB INTRONS AND THEIR CONTRIBUTION TO THE DIVERSIFICATION OF HLA.
          Katja Kotsch, Jenny Wehling, Raeiner  Blasczyk .  Department of Hematology and Oncology, Blood Bank, Charité, Campus Virchow-Klinikum, Humboldt-University, Berlin, Germany.
          Little is known about the non-coding regions of the different DRB alleles which represent about 93% of the genes. In this study we have determined the sequence of the 3' 500 bp intron 1 fragment and 5' 700 bp fragment of intron 2 adjacent to exon 2 in all serologically defined HLA-DRB genes and their most frequent allelic subtypes.The intron sequences turned out to be highly polymorphic. Besides extensive homologies, numerous locus- and group-specific sites including stretches of nucleotide deletions could be identified. Beyond that, there are close relationships between the HLA-DRB1 alleles belonging to the same haplotype group. Nearly all serological groups display at least one unique sequence motif not shared by any other group. With a few exceptions, the variability is arrested on the level of the serological diversity. Similar to the class I introns, this variability was not characterized by random point mutations but by a highly systematic diversity reflecting the lineage-specific relationship of the HLA-DRB alleles. The striking conservation within each ancestral lineage suggests that point mutations have been negatively selected. This finding strongly supports that introns are not as much subjected to evolutionary diversification forces as exons and puts forward the insertional theory of introns attributing them a functional relevance. An alternative interpretation of the conserved intron motifs would be that DRB alleles are too young to display intronic diversification. These data will deliver further insights in evolutionary mechanisms involved in DRB diversification and might provide a helpful tool for interspecies comparative analysis. Based on the now available intron data, it is likely that any new allele will carry the intron sequence motifs described so far. Consequently, the conserved intronic diversity will provide the possibility to establish advanced DRB sequencing typing strategies which do not require regular updates when new alleles are discovered.