Characterization and investigation of single nucleotide polymorphisms and a novel TLR2 mutation in the human TLR2 gene

Abstract

In the innate immune system, TLR2 plays a central role for the response to a wide variety of microbial and endogenous danger signals. A considerable number of genetic polymorphisms within the human TLR2 gene have been reported in non-coding and coding sequences. Except for the Arg753Gln variant, however, their clinical relevance is unclear and the assessment of the effects of amino acid substitutions on receptor function is lacking. In the present study, we have characterized all known single nucleotide polymorphisms (SNPs) of TLR2 for their functional relevance in transiently transfected HEK293 cells subsequently exposed to a specific stimulus. Among the known non-synonymous SNPs in the TLR2 coding sequence, four SNPs (Thr411Ile, Tyr715stop, Tyr715Lys and Arg753Gln) were found to be functionally relevant in our experimental setting. In addition, we identified a new mutation Arg447stop leading to a premature stop codon in the extracellular portion of the receptor. TLR2-specific stimulation of whole blood from two heterozygote donors of this mutation resulted in a reduced secretion of pro-inflammatory cytokines. Finally, we tested the prevalence of these functional genetic variants in 169 healthy individuals of Caucasian origin for the mutations in the extracellular domain and 106 individuals for the mutations in the intracellular domain of the receptor. Except for 10 heterozygote donors of the Arg753Gln variant determined to be prevalent in 9.4% of the tested individuals, none of the other SNPs was found in this population.

TLR2 Mediates Gap Junctional Intercellular Communication through Connexin-43 in Intestinal Epithelial Barrier Injury

“…Stable transfectants were selected with 1 g/ml blasticidin and maintained on poly-D-lysine-coated tissue culture plates (BD Biosciences). Transfection efficiency of the different clones was comparable with previous findings from other cell types (31), as tested by Western blotting and immunohistochemistry with anti-HA (InvivoGen; Cell Signaling). Caco-2 controls (mock) were untransfected cells without any exogenous DNA.…”

Genetic Variants of Toll-Like Receptor 2 and 5, Helicobacter Pylori Infection, and Risk of Gastric Cancer and Its Precursors in a Chinese Population

“…The TLR2 gene, mapped to chromosome 4q32, consists of 3 exons. Three major functional polymorphisms Pro631His, Arg677Trp, and Arg753Gln have been described in the Caucasian population (25,38,39). However, these SNPs are absent in our screening population, consistent with the HapMap database and another report (28).…”

Therapeutic Targeting of Toll-Like Receptors for Infectious and Inflammatory Diseases and Cancer

“…Mice without TLR2 are hyporesponsive to Gram-positive bacterial cell wall components . A number of SNPs in TLR2 have been reported in the extracellular domain [R753Q , Y715K, and Y715stop (Merx et al, 2007)] and the cytoplasmic domain [P631H (Smirnova et al, 2003)]; although the extracellular domain mutations result in decreased activity of TLR2 to Gram-positive bacterial ligands Schröder et al, 2003), the genetic evidence linking these SNPs with a susceptibility to Gram-positive infections is unclear (Schröder and Schumann, 2005). Twelve SNPs in TLR1 [of these, S248N, H305L, P315L (Johnson et al, 2007), and I602S (Johnson et al, 2007) have defective signaling] and 14 SNPs in TLR6 (Johnson et al, 2007) have also been identified.…”