Isolation of RNA aptamers against human Toll-like receptor 3 ectodomain.
Toll-like receptor 3 (TLR3) detects double-stranded RNA (dsRNA) known as a universal viral molecular pattern and activates the antiviral immune response. While TLR3 preferentially recognizes polyriboinosine-polyribocytidylic acid (poly (I:C)), a sequence-specific dsRNA has not yet been shown to activate TLR3. To determine whether TLR3 preferentially recognizes some specific sequence that acts on the signaling pathway of TLR3, in vitro selection against human TLR3 ectodomain (TLR3 ECD) was performed. After seventh selection cycle, two major classes, Family-I and -II, were emerged from 64 clones with binding constants of about 3 nM. Although these aptamers bound to TLR3 ECD with high affinity in vitro, they did not have agonist and antagonist effects on TLR3 signaling in TLR3-transfected HEK293 cells. Further analyses of the structure/function relationship of these aptamers will be carried out by mutagenesis, RNase mapping and competition assay using poly (I:C).
Watanabe
Tomoya
T
Department of Material and Biological Chemistry, Faculty of Science, Yamagata University, Yamagata 990-8560, Japan.
Ito
Kiyoharu
K
Matsumoto
Misako
M
Seya
Tsukasa
T
Nishikawa
Satoshi
S
Hasegawa
Tsunemi
T
Fukuda
Kotaro
K
D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000302:isolation & purification; D002460:Cell Line; D017930:Genes, Reporter; D006801:Humans; D017434:Protein Structure, Tertiary; D052156:SELEX Aptamer Technique; D051196:Toll-Like Receptor 3 / Q000819:agonists / Q000037:antagonists & inhibitors / Q000737:chemistry*
D052157:Aptamers, Nucleotide; C495344:TLR3 protein, human; D051196:Toll-Like Receptor 3