Epigenetic regulation of differentialHLA-Aallelic expression levels

Abstract

MHC class I expression levels influence the strength of immune responses and represent another variable in determining outcome to disease beyond peptide binding alone. Identification of the HLA loci that vary in allelic expression levels and delineating the mechanism responsible for expression variation may provide the opportunity to modify their expression therapeutically. We have examined the expression levels of allelic lineages at the HLA-A locus in a sample of 216 European Americans using a real-time polymerase chain reaction assay, which amplifies all HLA-A lineages specifically with equal efficiency, and observed a gradient of expression that associates with HLA-A allelic lineage (R = 0.6, P = 5 × 10(-25)). DNA methylation of the HLA-A gene appears to contribute to the variation in HLA-A mRNA expression levels, as a significant inverse correlation was observed between HLA-A mRNA expression levels in untreated cells and the degree to which expression is increased after treatment of the cells with a DNA methyltransferase inhibitor (R = 0.6, P = 2.8 × 10(-6)). Further, deep-sequencing and immunoprecipitation assays revealed allelic lineage-specific methylation patterns within the HLA-A promoter region where increased DNA methylation levels correlated significantly with reduced HLA-A expression levels (R = 0.89, P = 3.7 × 10(-9)). These data demonstrate HLA-A allelic lineage-specific variation in expression levels, and DNA methylation as a likely factor in contributing to this variation.

Posttranscriptional Regulation of HLA-A Protein Expression by Alternative Polyadenylation Signals Involving the RNA-Binding Protein Syncrip

“…Briefly, reverse transcription was performed with 900ng of total RNA using the high capacity RNA to cDNA kit (Applied Bioscience) in a volume of 10µl. HLA-A coding region [amplified using previously published primers (3)], HLA-A 3’UTR and GAPDH transcripts were amplified (Primer sequences in Table S1) by SYBR green qPCR using the threshold cycle (C T ) method(5) in a Viia7 machine (Applied Bioscience). Each qPCR reaction included 6µl of power SYBR green PCR mastermix (Applied Biosystems), 200nM primers that specifically amplified the gene of interest ( HLA-A ) or the housekeeping gene ( GAPDH ), and 2 µl of cDNA (1:20 dilution) in a total volume of 12 µl.…”

“…The mRNA expression levels of HLA-A alleles were recently shown to vary in an allele-dependent manner as a function of the degree of methylation in the promoter region of each allele (3). Thus, epigenetic mechanisms account for a portion of the differential mRNA expression patterns across HLA-A alleles.…”

Sequence and Phylogenetic Analysis of the Untranslated Promoter Regions for HLA Class I Genes

“…Binding sites for regulatory proteins further upstream from the core promoter have also been identified (26, 27) and it is likely that others exist, some of which may be polymorphic. Indeed, genetic variants and epigenetic modifications located within the 3´ and 5´ untranslated regions (UTR) (6, 13,25, 28, 29), as well as the coding region (14), have been shown to alter HLA allele expression levels, but so far, these mechanisms account for only a small percentage of the expression variation found within the respective genes.…”

Minimizing the risk of allo-sensitization to optimize the benefit of allogeneic cardiac-derived stem/progenitor cells

“…In healthy individuals, the expression levels of HLA-I and -II are allele dependent. This differential expression of HLA antigens is of great relevance in clinical setting47484950. Thus, beside DSA-HLA-I binding strength, their risk to hCPC should probably be viewed in the context of the HLA alleles of therapeutic cells.…”