Prediction of Altered 3′- UTR miRNA-Binding Sites from RNA-Seq Data: The Swine Leukocyte Antigen Complex (SLA) as a Model Region

Abstract

The SLA (swine leukocyte antigen, MHC: SLA) genes are the most important determinants of immune, infectious disease and vaccine response in pigs; several genetic associations with immunity and swine production traits have been reported. However, most of the current knowledge on SLA is limited to gene coding regions. MicroRNAs (miRNAs) are small molecules that post-transcriptionally regulate the expression of a large number of protein-coding genes in metazoans, and are suggested to play important roles in fine-tuning immune mechanisms and disease responses. Polymorphisms in either miRNAs or their gene targets may have a significant impact on gene expression by abolishing, weakening or creating miRNA target sites, possibly leading to phenotypic variation. We explored the impact of variants in the 3′-UTR miRNA target sites of genes within the whole SLA region. The combined predictions by TargetScan, PACMIT and TargetSpy, based on different biological parameters, empowered the identification of miRNA target sites and the discovery of polymorphic miRNA target sites (poly-miRTSs). Predictions for three SLA genes characterized by a different range of sequence variation provided proof of principle for the analysis of poly-miRTSs from a total of 144 M RNA-Seq reads collected from different porcine tissues. Twenty-four novel SNPs were predicted to affect miRNA-binding sites in 19 genes of the SLA region. Seven of these genes (SLA-1, SLA-6, SLA-DQA, SLA-DQB1, SLA-DOA, SLA-DOB and TAP1) are linked to antigen processing and presentation functions, which is reminiscent of associations with disease traits reported for altered miRNA binding to MHC genes in humans. An inverse correlation in expression levels was demonstrated between miRNAs and co-expressed SLA targets by exploiting a published dataset (RNA-Seq and small RNA-Seq) of three porcine tissues. Our results support the resource value of RNA-Seq collections to identify SNPs that may lead to altered miRNA regulation patterns.

The impact of breed and tissue compartment on the response of pig macrophages to lipopolysaccharide

“…Amongst the differences was the apparent absence of expression of SLA6 (Figure 8) and highly variable expression of SLA-DOA. These differences might be associated with polymorphic variation in miRNA recognition sites reported elsewhere [31]. Significant levels of protein-coding polymorphism have already been reported amongst pig pattern intracellular and extracellular recognition receptors [19,32].…”

Identification and annotation of conserved promoters and macrophage-expressed genes in the pig genome

“…RNA-Seq libraries were prepared from a male Large White x Landrace pig (about 7 weeks old). Unstranded 35 bp paired-end RNA-Seq libraries from alveolar macrophages (AM) were generated following the protocol described in [ 51 ]. Unstranded 57 bp paired-end RNA-Seq libraries from bone-marrow-derived macrophages (BMDM) were generated following the protocol described in [ 52 ].…”

PCK1 is negatively regulated by bta-miR-26a, and a single-nucleotide polymorphism in the 3′ untranslated region is involved in semen quality and longevity of Holstein bulls

“…miRNAs down‐regulate gene expression by binding to the 3′UTR of their target mRNAs, thus reducing transcript stability and translation. Polymorphisms in the miRNA or its binding sequence in its target can impact gene expression by abolishing, weakening, or creating miRNA‐binding sites, possibly resulting in phenotypic differences (Endale Ahanda et al, ). Given the previous prediction that bta‐miR‐26a binds this transcript and the relationship between SNPs in the PCK1 3′UTR and bull reproductive traits, we asked if bta‐miR‐26a can directly alter the expression of the PCK1 variants.…”