OR20 Investigating the microrna profile of B cell lines for their differential effect on haplotype specific DPB1 gene expression

Abstract

Aim SNP rs9277534 in the 3’-UTR region of the HLA-DPB1 gene is associated with differential expression of DP molecule on the cell surface of lymphocytes. It appears that this differential expression of DP influences risk of GVHD in unrelated donor hematopoietic cell transplantation and course of hepatitis B infection. We aim to uncover mechanisms for such alterations in protein expression by identifying haplotype-specific microRNAs (miRNAs) and their differential in silico targeting patterns that occur at polymorphic sites, with eventual experimental confirmation. Methods Novel and miRBase-annotated miRNAs expressed in COX (high DP expression) and PGF (low DP expression) B-lymphoblastoid cell lines (BLCL) were characterized using miRDeep2. Higher-confidence miRNAs were obtained by requiring > 80% estimated true positive probability, RANDFOLD p ⩽ 0.05, and removal of entries overlapping non-miRNA Rfam elements. A miRNA target prediction scheme utilizing complementary TargetScan v7 (TS) and RNA22v2 (R22) was employed, using the following relatively stringent parameter settings to identify the highest-precision targets: context++ score Results COX and PGF BLCLs demonstrated differential miRNA expression and predicted targeting at polymorphic sites of their respective DPB1 genes. Differential predicted targets were identified in exon 2, introns 1, 2, and 3 and the 3’-UTR of the two DPB1 sequences. These identified miRNAs have the potential to explain altered protein expression levels via putative cytoplasmic or nuclear miRNA effects. rs9277534 is not included among the high-confidence targeted sites; however, if thresholds are relaxed, then that region is differentially targeted as well. Conclusions We have identified miRNAs with differential expression in COX and PGF BLCLs, which demonstrate unique predicted targeting patterns at polymorphic sites of their respective HLA-DPB1 alleles. This catalog of miRNAs can now be tested in further experiments to identify true mediators of HLA-DPB1 expression and help to identify actual causative polymorphisms in such regions that involve extensive and confounding linkage disequilibrium.