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Functional analysis of differences in transcriptional activity conferred by genetic variants in the 5′ flanking region of the IL12RB2 gene

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Abstract

Interleukin 12 receptor β chain (IL12RB2) is a crucial regulatory factor involved in cell-mediated immune responses, and genetic variants of the gene encoding IL12RB2 are associated with susceptibility to various immune-related diseases. We previously demonstrated that haplotypes with single nucleotide polymorphisms (SNPs) in the 5′ flanking region of IL12RB2, including −1035A>G (rs3762315) and −1023A>G (rs3762316), affect the expression of IL12RB2, thereby altering susceptibility to leprosy and periodontal diseases. In the present study, we identified transcription factors associated with the haplotype-specific transcriptional activity of IL12RB2 in T cells and NK cells. The −1023G polymorphism was found to create a consensus binding site for the transcription factor activating protein (AP)-1, and enzyme-linked immunosorbent assay (ELISA)-based binding assays showed that these SNPs enhanced AP-1 binding to this region. In reporter assays, suppression of JunB expression using siRNA eliminated differences in the −1035G/−1023G and −1035A/−1023A regions containing IL12RB2 promoter activity in Jurkat T cells and NK3.3 cells. These results suggested that the −1035/−1023 polymorphisms created differential binding affinities for JunB that could lead to differential IL12RB2 expression. Moreover, the −1035G and −1035A alleles formed binding sites for GATA-3 and myocyte enhancer factor-2 (MEF-2), respectively. Our data indicated that in addition to JunB, the SNP at −1035/−1023 influenced GATA-3 and MEF-2 binding affinity, potentially altering IL12RB2 transcriptional activity. These findings confirm the effects of rs3762315 and rs3762316 on IL12RB2 transcription. These genetic variants may alter cellular activation of T cells and NK cells and modify cell-mediated immune responses.

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Abbreviations

Ab:

Antibody

AP-1:

Activating protein-1

ELISA:

Enzyme-linked immunosorbent assay

HRP:

Horseradish peroxidase

IFN:

Interferon

IL:

Interleukin

IL12RB2:

Interleukin 12 receptor β chain

MEF:

Myocyte enhancer factor

OD:

Optical density

rhIL:

Recombinant human interleukin

siRNA:

Small interfering RNA

SNP:

Single nucleotide polymorphism

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Acknowledgments

This study was supported, in part, by Grants-in-Aid for Scientific Research (C) (nos. 26463150 and 26463151) from the Japan Society for the Promotion of Science and Grants-in-Aid for Researchers, Hyogo College of Medicine, 2013.

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Authors and Affiliations

  1. Department of Pathology, Hyogo College of Medicine, Nishinomiya, Japan

    Nahoko Kato-Kogoe, Hideki Ohyama, Koji Yamanegi, Naoko Yamada, Masaki Hata, Hiroshi Nishiura, Nobuyuki Terada & Keiji Nakasho

  2. Department of Biochemistry and Molecular Biology, Nihon University School of Dentistry at Matsudo, Matsudo, Japan

    Soichiro Okano & Yoshimitsu Abiko

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  1. Nahoko Kato-Kogoe
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  2. Hideki Ohyama
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  9. Nobuyuki Terada
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  10. Keiji Nakasho
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Correspondence to Nahoko Kato-Kogoe.

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Kato-Kogoe, N., Ohyama, H., Okano, S. et al. Functional analysis of differences in transcriptional activity conferred by genetic variants in the 5′ flanking region of the IL12RB2 gene. Immunogenetics 68, 55–65 (2016). https://doi.org/10.1007/s00251-015-0882-x

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  • DOI: https://doi.org/10.1007/s00251-015-0882-x

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