THE NATURE OF POLYMORPHISM OF THE HLA CLASS-I NONCODING REGIONS AND THEIR CONTRIBUTION TO THE DIVERSIFICATION OF HLA

The sequence database of HLA class I genes is mainly derived from mRNA analysis. Little is known about the non-coding sequences of the diffe rent class I alleles. In this study we have determined the sequence of the Ist through 3rd introns of the majority of HLA-A and -B alleles. The few published sequences emerged to contain substantial errors. The introns turned out to be highly polymorphic with a variability of 14. 6% in the Ist intron decreasing to 6.2% in the 3rd intron. Against all expectations, this variability is not characterised by random point m utations but by a highly systematic diversity reflecting the ancestral relationship of the HLA alleles. The variability is arrested on the l evel of the serological diversity. The striking conservation within ea ch ancestral lineage suggests that point mutations have been negativel y selected. This finding could be explained by the evolutionary pressu re on base order, promoting the potential to extrude single-strand ste m-loops from supercoiled duplex DNA, which is believed to be important for recombination. Moreover, the GC content was found to be as high a s 78% in the ist and 2nd introns and 55% in the 3rd intron. These CpG islands are directly involved in the exchange of short stretches of DN A in unequal crossing-over events. Additionally, conversion between di fferent class I sequences is Facilitated by regions of strong homology , stabilizing the pairing of variable regions. All these observations indicate the potential of a substantial contribution of introns to the recombinational activity of class I genes. The exclusive clustering o f CpG islands in the Ist and 2nd introns restricts the gene conversion events to the regions of the 2nd and 3rd exons and therefore protects the conservation of the 5' flanking region and the 3' part of the gen e. Since there are less diversification forces acting on introns they may be more conserved in a trans-species manner than exons. Therefore, they could provide the answer for the controversy regarding intra-or trans-species evolution.