The small GTPase Arf6 is essential for TLR4 signaling in multiple (path)ways

The human body’s first line of defense against invading pathogens is the innate immune system. This defense mechanism uses pattern recognition receptors (PRRs) that recognize pathogen-associated molecular patterns (PAMPs). One family of PRRs, is the Toll-like receptor (TLR) family consisting of 10 human members which all recognize different PAMPs. Upon detection of their ligands, TLRs initiate an intracellular signaling cascade resulting in either NF-κB activation and pro-inflammatory cytokine production (the MyD88-dependent pathway) or interferon-regulatory factor (IRF) activation and interferon production (the MyD88-independent pathway). TLR4 is a unique member of the TLR family as it is able to signal via both the MyD88-dependent and MyD88-independent pathway.Binding of lipopolysaccharides (LPS) to TLR4, results in the intracellular recruitment of the TLR4 adaptor proteins Mal and MyD88, initiating the MyD88-dependent signaling cascade. Upon endocytosis of the activated receptor, the adaptors Tram and Trif are recruited activating the MyD88-independent pathway. A tight regulation of activation of these pathways is required in order to prevent chronic inflammation and several associated diseases.The small GTPase ADP-ribosylation factor 6 (Arf6) was implicated in TLR4 signaling as a regulator of the localization of the adaptor protein Mal, hence controlling TLR4 signal transduction. However, later findings contradicted this. In this PhD, we clarified the role of Arf6 in TLR4 signaling. We showed that Arf6 is able to function independently of Mal upon LPS stimulation. This lead to the discovery of a novel function of Arf6 as a regulator of LPS-induced transport of the adaptor protein Tram to the endocytic recycling compartment. Via this mechanism, Arf6 controls the MyD88-independent TLR4 signaling pathway and subsequent interferon production. Next to its role in the MyD88-independent pathway, Arf6 is essential for the initial steps of the TLR4 MyD88-dependent signaling cascade resulting in NF-κB activation and pro-inflammatory cytokine production. We show for the first time that Arf6 is activated immediately upon LPS stimulation.These results demonstrate that Arf6 is an essential regulator of TLR4 signal transduction. Further research on the effector proteins, the GTPase-activating proteins (GAPs; deactivators of Arf6) and guanine nucleotide exchange factors (GEFs; Arf6 activators) that regulate/are regulated by Arf6 in its TLR4-specific functions might provide valuable information to develop new therapeutics targeting dysregulated TLR4 processes.Public defense: 2014-06-27The human body’s first line of defense against invading pathogens is the innate immune system. This defense mechanism uses pattern recognition receptors (PRRs) that recognize pathogen-associated molecular patterns (PAMPs). One family of PRRs, is the Toll-like receptor (TLR) family consisting of 10 human members which all recognize different PAMPs. Upon detection of their ligands, TLRs initiate an intracellular signaling cascade resulting in either NF-κB activation and pro-inflammatory cytokine production (the MyD88-dependent pathway) or interferon-regulatory factor (IRF) activation and interferon production (the MyD88-independent pathway). TLR4 is a unique member of the TLR family as it is able to signal via both the MyD88-dependent and MyD88-independent pathway.Binding of lipopolysaccharides (LPS) to TLR4, results in the intracellular recruitment of the TLR4 adaptor proteins Mal and MyD88, initiating the MyD88-dependent signaling cascade. Upon endocytosis of the activated receptor, the adaptors Tram and Trif are recruited activating the MyD88-independent pathway. A tight regulation of activation of these pathways is required in order to prevent chronic inflammation and several associated diseases.The small GTPase ADP-ribosylation factor 6 (Arf6) was implicated in TLR4 signaling as a regulator of the localization of the adaptor protein Mal, hence controlling TLR4 signal transduction. However, later findings contradicted this. In this PhD, we clarified the role of Arf6 in TLR4 signaling. We showed that Arf6 is able to function independently of Mal upon LPS stimulation. This lead to the discovery of a novel function of Arf6 as a regulator of LPS-induced transport of the adaptor protein Tram to the endocytic recycling compartment. Via this mechanism, Arf6 controls the MyD88-independent TLR4 signaling pathway and subsequent interferon production. Next to its role in the MyD88-independent pathway, Arf6 is essential for the initial steps of the TLR4 MyD88-dependent signaling cascade resulting in NF-κB activation and pro-inflammatory cytokine production. We show for the first time that Arf6 is activated immediately upon LPS stimulation.These results demonstrate that Arf6 is an essential regulator of TLR4 signal transduction. Further research on the effector proteins, the GTPase-activating proteins (GAPs; deactivators of Arf6) and guanine nucleotide exchange factors (GEFs; Arf6 activators) that regulate/are regulated by Arf6 in its TLR4-specific functions might provide valuable information to develop new therapeutics targeting dysregulated TLR4 processes.D