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A functional polymorphism of PON1 interferes with microRNA binding to increase the risk of ischemic stroke and carotid atherosclerosis

  • Mu-En Liu
    Affiliations
    Department of Medical Genetics, Kaohsiung Medical University, Kaohsiung, Taiwan

    Department of Psychiatry, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan
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  • Yi-Chu Liao
    Affiliations
    Department of Neurology, Taichung Veterans General Hospital, Taichung, Taiwan

    Department of Neurology, National Yang-Ming University School of Medicine, Taipei, Taiwan
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  • Ruey-Tay Lin
    Affiliations
    Department of Neurology, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Department of Neurology, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
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  • Yung-Song Wang
    Affiliations
    Department of Medical Genetics, Kaohsiung Medical University, Kaohsiung, Taiwan

    Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
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  • Edward Hsi
    Affiliations
    Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
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  • Hsiu-Fen Lin
    Affiliations
    Department of Neurology, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan

    Department of Neurology, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan
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  • Ku-Chung Chen
    Affiliations
    Department of Medical Genetics, Kaohsiung Medical University, Kaohsiung, Taiwan

    Department of Biochemistry, College of Medicine, Taipei Medical University, Taipei, Taiwan
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  • Suh-Hang H. Juo
    Correspondence
    Corresponding author. Department of Medical Genetics, Kaohsiung Medical University, No. 100, TzYou First Road, Kaohsiung 80708, Taiwan. Tel.: +886 7 312 1101x6470; fax: +886 7 321 3931.
    Affiliations
    Department of Medical Genetics, Kaohsiung Medical University, Kaohsiung, Taiwan

    Department of Medical Research, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
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      Abstract

      Objective

      Single nucleotide polymorphisms (SNPs) located at microRNA (miRNA) binding sites (miR-SNPs) can affect the expression of genes. This study aimed to identify the miR-SNPs associated with atherosclerosis and stroke.

      Methods

      Patients with ischemic stroke (n = 657) and stroke- and myocardial infarction-free volunteers (n = 1571) were enrolled. The carotid intima-media thickness (IMT) was measured in the control participants. Seventy-nine stroke susceptibility genes were initially selected and 13 genes were predicted to have miR-SNPs at their 3′ untranslated regions (3′UTR). The miRNA arrays were used to further identify potential miR-SNPs. The miR-SNP rs3735590 at the paraoxonase 1 (PON1) gene was finally selected and its associations with stroke and carotid IMT were evaluated. The 3′UTR reporter and SNP functional assays were then performed to validate the results.

      Results

      Compared with CC genotype, patients with CT or TT genotype at rs3735590 had lower risk of ischemic stroke (OR = 0.72, p = 0.036; OR = 0.83, p = 0.077, respectively). Among the healthy participants, the CT or TT genotype was associated with thinner IMT in the internal carotid arteries in comparison with CC genotype (β = −0.76, p = 0.003; β = −0.022, p = 0.452, respectively). Our findings suggested that the minor allele T had a protective effect on atherosclerosis. Results from 3′UTR reporter assays showed that PON1 is a direct target gene of miR-616. In plasmid constructs carrying the risk allele C at rs3735590, miR-616 inhibited the genetic expression of PON1. However, substitution of C by T at rs3735590 reduced the miR-616 binding affinity, leading to overexpression of the PON1 gene.

      Conclusion

      Our study is the first to show that the miR-SNP at PON1 could affect genetic expression and is associated with an elevated risk for ischemic stroke and subclinical atherosclerosis.

      Highlights

      • T allele carriers of PON1 rs3735590 had lower risk of ischemic stroke.
      • T allele carriers had thinner intima-media thickness of internal carotid arteries.
      • PON1 is a novel direct target gene of miR-616 by 3′UTR reporter assays.
      • The binding ability of miR-616 and PON1 gene expression is regulated by SNP rs3735590.

      Keywords

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      Linked Article

      • MicroRNA-616 and atherosclerosis – A commentary on the paper by Liu et al.
        AtherosclerosisVol. 228Issue 1
        • Preview
          MicroRNAs (miRNA, miR) in cardiovascular diseases are a constantly growing field. As individual microRNAs are predicted to regulate the expression of hundreds of genes, and a single gene can be targeted by numerous microRNAs, microRNA profiling can rarely indicate the mechanism through which microRNAs affect the disease. In their paper “A functional polymorphism of PON1 interferes with microRNA binding to increase the risk of ischemic stroke and carotid atherosclerosis”, Liu et al. have shown a decreased binding affinity of miR-616 to the 3′-UTR as well as an increased paraoxonase 1 (PON1) expression in a plasmid construct carrying the T allele of the single-nucleotide polymorphism (SNP) rs3735590.
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