title: Antagonistic RNA aptamer specific to a heterodimeric form of human interleukin-17A/F.
abstract: Interleukin-17 (IL-17) is a pro-inflammatory cytokine produced primarily by a subset of CD4(+)T cells, called Th17 cells, that is involved in host defense, inflammation and autoimmune disorders. The two most structurally related IL-17 family members, IL-17A and IL-17F, form homodimeric (IL-17A/A, IL-17F/F) and heterodimeric (IL-17A/F) complexes. Although the biological significance of IL-17A and IL-17F have been investigated using respective antibodies or gene knockout mice, the functional study of IL-17A/F heterodimeric form has been hampered by the lack of an inhibitory tool specific to IL-17A/F. In this study, we aimed to develop an RNA aptamer that specifically inhibits IL-17A/F. Aptamers are short single-stranded nucleic acid sequences that are selected in vitro based on their high affinity to a target molecule. One selected aptamer against human IL-17A/F, AptAF42, was isolated by repeated cycles of selection and counterselection against heterodimeric and homodimeric complexes, respectively. Thus, AptAF42 bound IL-17A/F but not IL-17A/A or IL-17F/F. The optimized derivative, AptAF42dope1, blocked the binding of IL-17A/F, but not of IL-17A/A or IL-17F/F, to the IL-17 receptor in the surface plasmon resonance assay in vitro. Consistently, AptAF42dope1 blocked cytokine GRO-α production induced by IL-17A/F, but not by IL-17A/A or IL-17F/F, in human cells. An RNA footprinting assay using ribonucleases against AptAF42dope1 in the presence or absence of IL-17A/F revealed that part of the predicted secondary structure fluctuates between alternate forms and that AptAF42dope1 is globally protected from ribonuclease cleavage by IL-17A/F. These results suggest that the selected aptamer recognizes a global conformation specified by the heterodimeric surface of IL-17A/F.
authors.affiliation: Department of Basic Medical Sciences, Institute of Medical Science, The University of Tokyo, Minato-ku, Tokyo, Japan.
authors.forename: Hironori
authors.initials: H
authors.lastname: Adachi
authors.forename: Akira
authors.initials: A
authors.lastname: Ishiguro
authors.forename: Michiaki
authors.initials: M
authors.lastname: Hamada
authors.forename: Eri
authors.initials: E
authors.lastname: Sakota
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Asai
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Nakamura
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000378:metabolism* / Q000494:pharmacology; D001483:Base Sequence; D001667:Binding, Competitive; D003198:Computer Simulation; D006801:Humans; D020381:Interleukin-17 / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D055503:Protein Multimerization; D053722:Receptors, Interleukin-17 / Q000378:metabolism; D012260:Ribonucleases / Q000378:metabolism; D020349:Surface Plasmon Resonance
chemical_list: D052157:Aptamers, Nucleotide; D020381:Interleukin-17; D053722:Receptors, Interleukin-17; D012260:Ribonucleases