A unique second donor splice site in the intron 5 sequence of the HLA-A*11alleles results in a class I transcript encoding a molecule with an elongated cytoplasmic domain

Full-length cDNA sequencing of the A*1103 allele revealed an insertion of 1 8 bp between exon 5 and 6. We hypothesized that this could be the result of alternative splicing. Sequencing of intron-5 of A*1101, *1102 and *1103 al leles demonstrated that this 18-bp insertion consisted of the 5'-end of int ron 5, concluded by a second in-frame donor splice site. Alignment of the 5 '-end intron 5 sequence of A*1101-3 with that of A*0101, *0201 and 0301 rev ealed a unique polymorphism at position 17 of the intron (A to G) that crea ted this second donor splice site. To exclude the possibility of an Epstein -Barr virus (EBV)-induced event, reverse transcriptase-polymerase chain rea ction (RT-PCR) analysis was performed on both peripheral blood mononuclear cells (PBMC) and EBV-transformed b-LcL's of several A*11-positive individua ls, using primers spanning exons 5 and 6, Without exception, both cell type s revealed two products for A*11, Densitometric analysis using EBV-transfor med b-LcL's and PBMC indicated a ratio of approximately 4:1 in favor of the alternative splice product. Notably, except for the A*11's none of the oth er A-locus alleles yields this alternative splice product. Translation of t his product will result in a protein that has an additional 6 amino acids i n its cytoplasmic domain. This introduces a negative charge just behind the basic anchor residues of the cytoplasmic segment and results in the loss o f the single potential protein kinase C phosphorylation site.