In vivo analysis of HLA-DQ gene expression in heterozygous cell lines

Abstract

Regulation of HLA-DQ gene transcription is a complex phenomenon because the allelic polymorphism associated with these genes and their promoters is a putative source of differential allele expression. Both transcriptional and post-transcriptional regulation could account for the density of the molecules expressed at the cell surface and then for the specificity of the immune response. Different methods have been developed to evaluate the functional consequences of this polymorphism, but at present no universal method allows measurement of either the steady-state level or the half-life time of mRNA species of both DQA1 and DQB1 polymorphic genes in heterozygous cell lines. Here, we propose a potent method, based on relative quantification of reverse transcriptase-polymerase chain reaction products, which analyzes the differential expression of all DQA1 or DQB1 allele combinations. This method is used to analyze the differential expression of HLA-DQB1*020110402 alleles in the human heterozygous lymphoblastoid B-cell line. Nucleotidic sequences of the proximal upstream regulatory region of these alleles exhibit significant differences. We show that the DQB1*0402 promoter is able to mediate a transcription strength twice as efficiently as *0201. In addition, the *0402 mRNA steady-state level is also governed by a remarkable post-transcriptional regulation. Indeed, an important part (20%) of the *0402 primary transcript is derived by alternative splicing in a short mRNA translated into a nonfunctional protein. Despite their variable sequence and length, no difference in the half-life of DQB1*0201 and both DQB*0402 mRNAs was observed in B-lymphoblastoid cells. The implications of these findings are discussed.