Abstract
Toll-like receptors (TLR) are highly conserved pattern recognition receptors of the innate immune system. Toll-like receptor 2 (TLR2) recognizes bacterial lipopeptides in a heterodimeric complex with TLR6 or TLR1, thereby discriminating between di- or triacylated lipopeptides, respectively. Previously, we found that HEK293 cells transfected with bovine TLR2 (boTLR2) were able to respond to diacylated lipopeptides but did not recognize triacylated lipopeptides, even after cotransfection with the so far published sequence of boTLR1. In this study we now could show that primary bovine cells were in general able to detect triacylated lipopetides. A closer investigation of the boTLR1 gene locus revealed an additional ATG 195 base pairs upstream from the published start codon. Its transcription would result in an N-terminus with high identity to human and murine TLR1 (huTLR1, muTLR1). Cloning and cotransfection of this longer boTLR1 with boTLR2 now resulted in the recognition of triacylated lipopeptides by HEK293 cells, thereby resembling the ex vivo observation. Analysis of the structure-activity relationship showed that the ester-bound acid chains of these lipopeptides need to consist of at least 12 carbon atoms to activate the bovine heterodimer showing similarity to the recognition by huTLR2/huTLR1. In contrast, HEK293 cell cotransfected with muTLR2 and muTLR1 could already be activated by lipopeptides with shorter fatty acids of only 6 carbon atoms. Thus, our data indicate that the additional N-terminal nucleotides belong to the full length and functionally active boTLR1 (boTLR1-fl) which participates in a species-specific recognition of bacterial lipopeptides.
Highlights
A prerequisite for the induction of an immune response against invading pathogens is the recognition of their presence
When transfected with bovine TLR2 (boTLR2) only diacylated lipopeptides were recognized indicating a heterodimerization with native human TLR6 but not with human TLR1
In our previous study we could show, that HEK293 cells stably transfected with boTLR2 were able to respond to Staphylococcus aureus, heat-killed Listeria monocytogenes (HKLM), Escherichia coli causing mastitis and to different diacylated lipopeptides [9]
Summary
A prerequisite for the induction of an immune response against invading pathogens is the recognition of their presence. With the exception of TLR10, 12 and 13 natural ligands are known for all TLR allowing the immune system to detect most kinds of invading microorganisms [2, 22]. The TIR domain is highly conserved between TLR and different species and mediates the induction of signal transduction cascades via MyD88 and/or as found with TLR3 and TLR4 via TRIF [7]. Ligand binding occurs via the LRR of the ectodomain. For this reason this domain highly varies between different TLR but shares great similarity between the same TLR of different species [24]. There are some species-specific differences in the ligand recognition [31]
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