Functional relevance of the IRF-1 promoter polymorphism rs2549009 on transcriptional activity in a native genomic environment

Abstract

Interferon regulatory factor-1 (IRF-1), a transcription regulator involved both in inducing and in mediating the effects of interferon, is encoded by a highly polymorphic gene in different ethnic populations. Some of these genetic variations have been described to be associated to disease traits in hepatitis C virus and in human immunodeficiency virus infection, including one single-nucleotide polymorphism rs2549009 within the promoter region. This study aimed at investigating the functional relevance of rs2549009 on IRF-1 transcriptional activity in peripheral blood mononuclear cells in its natural genomic environment. Haplotype-specific chromatin immunoprecipitation using antibodies directed against both the transcriptionally inactive and active RNA polymerase II (RNAPII) and allele-specific transcript quantification techniques were applied to ex vivo-derived samples from healthy heterozygous donors. Inactive serine 5 phosphorylated RNAPII was found to be preferentially bound to the rs2549009 A allele in all donors investigated. Active serine 2 phosphorylated (ser2-P) RNAPII, in contrast, was found to be precipitable, depending on the donor, preferentially either with the A or the G promoter variants or without any preference. The ratio of rs2549009 A/G promoter variants engaged by ser2-P RNAPII was closely related to the relative frequency of the respective IRF-1 transcripts, and relative allelic expression was found to be associated to total IRF-1 gene expression. These results provide evidence for a bidirectional IRF-1 gene expression imbalance that appears not to be solely controlled by rs2549009 in cis and may rely on a yet unidentified variant or haplotype or on environmental control in trans.

Interaction between genetic and epigenetic variation defines gene expression patterns at the asthma-associated locus 17q12-q21 in lymphoblastoid cell lines

“…C is -regulatory allelic effects may arise from allele-specific differences in transcription factor binding and enhancer activity (Agueda et al 2011; Bickel et al 2011; Colombo et al 2011; Harmon et al 2010; Mertens et al 2010). Transcription factor binding to a regulatory DNA element usually results in repositioning of nucleosomes.…”

The regulatory toll-like receptor 4 genetic polymorphism rs11536889 is associated with renal, coagulation and hepatic organ failure in sepsis patients

“…Peripheral blood monocytes (PBMCs) from approximately 30 ml of heparinized peripheral blood were isolated through Ficoll density gradient centrifugation according to standard procedures described previously [33]. Cell preparations were routinely assessed for viability (>95%) by trypan blue dye exclusion.…”

Exploring the Functional Relevance of Polymorphisms within the CD14 and IRF-1 Gene for Promoter Activity by Haplotype-Specific Chromatin Immunoprecipitation (HaploChIP)

“…Linear regression analysis of data derived from the two different read out assays revealed a close correlation with p = 0.003 and r = 0.71. These results provide first evidence for a bidirectional IRF-1 gene expression imbalance in PBMC (ASTQ experiments were performed by our research team members; Mertens et al, 2010).…”

“…A parallel analysis of total IRF-1 gene expression revealed that total IRF-1 gene expression was not significantly different when related to IRF-1 rs2549009 promoter genotypes (p = 0.2709) in 41 healthy Caucasians. A more detailed analysis within the group of 16 individuals heterozygous in IRF-1 rs2549009, however, revealed a close and positive relation (p = 0.0035, r = -0.68) between the relative amount of G allele transcripts and total IRF-1 mRNA levels in PBMC (these analyses were done in cooperation with our research team members; Mertens et al, 2010).…”

“…Two other SNPs at position -410 (IRF-1 rs2549007) and -415 (IRF-1 rs2706384) that have been described as in a strong LD with IRF-1 rs2549009 (Saito et al, 2001) are marked in dark blue. Intronic variations that have been described as associated with different virus diseases are given in green (Ball et al, 2007;Fortunato et al, 2008) (Modified from Mertens et al, 2010). Therefore, 13 healthy Caucasian blood donors were first of all genotyped at this variant position and IRF-1 rs2549009 heterozygous individuals (n=6) were used for the investigation of the allelic IRF-1 promoter activity.…”