The TLR4 D299G and T399I SNPs Are Constitutively Active to Up-Regulate Expression of Trif-Dependent Genes

Abstract

Dysregulated Toll-Like Receptor (TLR) signalling and genetic polymorphisms in these proteins are linked to many human diseases. We investigated TLR4 functional variants D299G and T399I to assess the impact on LPS-induced responsiveness in comparison to wild-type TLR4. The mechanism by which this occurs in unclear as these SNPs do not lie within the lipid A binding domain or dimerisation sites of the LPS-TLR4/MD2 receptor complexes. Transfection of TLR4D299G, TLR4T399I or TLR4D299G. T399I into HEK cells resulted in constitutive activation of an NF-κB reporter gene and a blunting of the LPS-induced reporter activation compared to WT-TLR4. Unstimulated human monocyte/macrophages, from patients with the D299G and T399I SNPs demonstrated a downregulation of many genes, particularly Tram/Trif signalling pathway constitutents compared to the TLR4 wild-type subjects supporting the concept of basal receptor activity. Monocyte/macrophages from carriers of the TLR4 D299G and T399I polymorphisms stimulated with LPS showed >6 fold lower levels of NF-κB and ∼12 fold higher IFN-β gene expression levels compared to wild-type subjects (P<0.05; MWU test) and dramatically altered resultant cytokine profiles. We conclude that these TLR4 SNPs affect constitutive receptor activity which impacts on the hosts ability to respond to LPS challenge leading to a dysregulated sub-optimal immune response to infection.

Single nucleotide polymorphisms of Toll-like receptors and susceptibility to infectious diseases

“…Additionally, they showed that many genes, particularly the TRAM/TRIF signalling pathway constituents, are down‐regulated in unstimulated human monocyte/macrophages from patients with the D299G and T399I SNPs, while upon LPS stimulation these cells display lower NF‐κB levels, higher IFN‐β gene expression levels and overall altered cytokine profiles, compared to the wild‐type receptor. Taken together, these data support that these common TLR‐4 SNPs affect constitutive receptor activity, thus impacting that ability of the host to respond to LPS challenge and a suboptimal immune response to infection .…”

Association of Helicobacter pylori infection with Toll‐like receptor‐4 Thr399Ile polymorphism increased the risk of peptic ulcer development in North of Iran

“…Yet, not all studies have confirmed the finding. For example, Georgina et al from America reported that Asp299Gly could be considered as a risk factor for non-cardia gastric carcinoma and its precursors (31).…”

For when bacterial infections persist: Toll-like receptor-inducible direct antimicrobial pathways in macrophages

“…For example, individuals carrying TLR4 Asp299Gly (A896G) and Thr399Ile (C1196T) variants have a higher prevalence of Gram‐negative infections and chronic periodontal disease . These mutations actually lead to constitutive basal activity of the receptor and a dampening of LPS‐inducible inflammatory responses, similar to the phenomenon of tolerisation . In general, SNPs in TLR‐related genes are often associated with hyporesponsiveness to activating stimuli, leading to less efficient immune responses and increased susceptibility to infection.…”