Phosphatidylinositol 4-Phosphate 5-Kinase α Facilitates Toll-like Receptor 4-mediated Microglial Inflammation through Regulation of the Toll/Interleukin-1 Receptor Domain-containing Adaptor Protein (TIRAP) Location*

Nguyen, Tu Thi Ngoc; Kim, Yong Min; Kim, T. Doohun; Le, Oanh Thi Tu; Kim, Jae Jin; Kang, Ho Chul; Hasegawa, Hiroshi; Kanaho, Yasunori; Jou, Ilo; Lee, Sang Yoon

doi:10.1074/jbc.m112.410126

Cited by 43 publications

(55 citation statements)

References 63 publications

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“…Phosphorylation of PI(4,5)P 2 to PI (3,4,5)P 3 by the p110δ isoform of phosphatidylinositol 3-kinase serves as a switch from MyD88-to TRIF-dependent signaling (Aksoy et al, 2012), and PI(4,5)P 2 hydrolysis can also contribute to this switch (Chiang et al, 2012). It has recently been reported that LPS induces an increase of the PI(4,5)P 2 level in microglia by activating PIP5K Iα (Nguyen et al, 2013).…”

Section: Introductionmentioning

confidence: 99%

LPS-induced clustering of CD14 triggers generation of PI(4,5)P2

Płóciennikowska

Zdioruk

Traczyk

et al. 2015

Journal of Cell Science

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Bacterial lipopolysaccharide (LPS) induces strong pro-inflammatory reactions after sequential binding to CD14 protein and TLR4 receptor. Here, we show that CD14 controls generation of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P 2 ] in response to LPS binding. In J774 cells and HEK293 cells expressing CD14 exposed to 10-100 ng/ml LPS, the level of PI(4,5)P 2 rose in a biphasic manner with peaks at 5-10 min and 60 min. After 5-10 min of LPS stimulation, CD14 underwent prominent clustering in the plasma membrane, accompanied by accumulation of PI(4,5)P 2 and type-I phosphatidylinositol 4-phosphate 5-kinase (PIP5K) isoforms Iα and Iγ (encoded by Pip5k1a and Pip5k1c, respectively) in the CD14 region. Clustering of CD14 with antibodies, without LPS and TLR4 participation, was sufficient to trigger PI(4,5)P 2 elevation. The newly generated PI(4,5)P 2 accumulated in rafts, which also accommodated CD14 and a large portion of PIP5K Iα and PIP5K Iγ. Silencing of PIP5K Iα and PIP5K Iγ, or application of drugs interfering with PI(4,5)P 2 synthesis and availability, abolished the LPS-induced PI(4,5)P 2 elevation and inhibited downstream pro-inflammatory reactions. Taken together, these data indicate that LPS induces clustering of CD14, which triggers PI(4,5)P 2 generation in rafts that is required for maximal pro-inflammatory signaling of TLR4.

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Section: Introductionmentioning

confidence: 99%

LPS-induced clustering of CD14 triggers generation of PI(4,5)P2

Płóciennikowska

Zdioruk

Traczyk

et al. 2015

Journal of Cell Science

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“…Indeed, association of TIRAP to membranes is transient during TLR-mediated signaling. LPS-mediated activation of TLRs induces dynamic changes in PtdIns(4,5)P 2 and TIRAP translocation to the plasma membrane, where TIRAP is transiently located at the plasma membrane and moves to the cytosol 30 min after LPS stimulation 12 . TIRAP meets the general features of a peripheral membrane protein.…”

Section: Discussionmentioning

confidence: 99%

“…Synthesis and turnover of plasma membrane PtdIns(4,5)P 2 influences TIRAP subcellular localization. Membrane binding of TIRAP is regulated by phosphatidylinositol-5 kinase (PI5K), an enzyme that generates intracellular PtdIns(4,5)P 2 and co-localizes with TIRAP at the plasma membrane 12 . Moreover, TIRAP interacts with PI3Ks, enzymes that convert PtdIns(4,5)P 2 into PtdIns(3,4,5)P 3 , impairing TIRAP's membrane targeting 13 .…”

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confidence: 99%

Membrane Targeting of TIRAP is Negatively Regulated by Phosphorylation in its Phosphoinositide-Binding Motif

Capelluto

Xiong

Zhao

et al. 2017

Biophysical Journal

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1Pathogen-activated Toll-like receptors (TLRs), such as TLR2 and TLR4, dimerize and move laterally across the plasma membrane to phosphatidylinositol (4,5)-bisphosphate-enriched domains. At these sites, TLRs interact with the TIR domain-containing adaptor protein (TIRAP), triggering a signaling cascade that leads to innate immune responses. Membrane recruitment of TIRAP is mediated by its phosphoinositide (PI)-binding motif (PBM). We show that TIRAP PBM transitions from a disordered to a helical conformation in the presence of either zwitterionic micelles or monodispersed PIs. TIRAP PBM bound PIs through basic and nonpolar residues with high affinity, favoring a more ordered structure. TIRAP is phosphorylated at Thr28 within its PBM, which leads to its ubiquitination and degradation. We demonstrate that phosphorylation distorts the helical structure of TIRAP PBM, reducing PI interactions and cell membrane targeting. Our study provides the basis for TIRAP membrane insertion and the mechanism by which it is removed from membranes to avoid sustained innate immune responses.The innate immune system is composed of germline-encoded receptor proteins that recognize invading pathogens by what is referred to as pathogen associated molecular patterns (PAMPs)1 . Toll-like receptors (TLRs) are the best-studied group of innate immune receptors that are able to recognize various PAMPs, such as bacterial lipopolysaccharides (LPS) or double stranded RNA from viruses 2 . Ligand-activated TLRs self-associate, triggering the recruitment of cytoplasmic adaptor proteins. Through their TIR domains, TLR2 and TLR4 interact with the TIR domain-containing adaptor protein (TIRAP; also known as MAL) and the myeloid differentiation primary response gene 88 (MyD88) [reviewed in ref. 3]. TIRAP contains an N-terminal phosphoinositide (PI)-binding domain (PBD) followed by a TIR domain. Plasma membrane localization of TIRAP depends on the presence of phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P 2 )-enriched regions 4 . TIRAP also serves as a bridge to recruit MyD88 through TIR-TIR domain interactions 5 . However, the presence of TIRAP at the plasma membrane is required even when there are low TLR ligand levels 6 . TIRAP's association triggers further recruitment of members of the IRAK family of kinases to promote formation of the myddosome, which in turn activates TRAF6 and NF-κ B nuclear translocation 7 . As a transcription factor, NF-κ B mediates pro-inflammatory and anti-microbial gene expression. The structure of the TIRAP TIR domain reveals two potential dimerization interfaces in a configuration that allows the two monomers of the N-terminal PBD to be oriented in the same direction, facilitating PtdIns(4,5)P 2 -mediated plasma membrane targeting 8,9 . Plasma membrane targeting of TIRAP is likely mediated by a short stretch of basic residues [amino acids 15-35; herein named PI-binding motif (PBM)] within the putative PBD 10 . Alanine mutagenesis of TIRAP residues Lys15, Lys16, Lys31, and Lys32 or hydrolysis of cellular PtdIns(4,5)P 2...

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“…The same group confirmed these initial observations by evidencing that a dominant-negative mutant of PIP5Ka impaired PIP2 production and the membrane recruitment of TIRAP in LPS-stimulated microglia. As a result, TLR4-associated signalling pathways and pro-inflammatory mediator production were both strongly inhibited [69]. Based on these observations, a role for PIP5K in regulating critical functions of macrophages, ranging from the ingestion and elimination of pathogens to the signals regulating the expression of genes involved in host defence from infection, can be envisaged (Fig.…”

Section: Macrophagesmentioning

confidence: 99%

The multifaceted role of PIP2 in leukocyte biology

Tuosto

Capuano

Muscolini

et al. 2015

Cell. Mol. Life Sci.

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Phosphatidylinositol 4,5-bisphosphate (PIP2) represents about 1 % of plasma membrane phospholipids and behaves as a pleiotropic regulator of a striking number of fundamental cellular processes. In recent years, an increasing body of literature has highlighted an essential role of PIP2 in multiple aspects of leukocyte biology. In this emerging picture, PIP2 is envisaged as a signalling intermediate itself and as a membrane-bound regulator and a scaffold of proteins with specific PIP2 binding domains. Indeed PIP2 plays a key role in several functions. These include directional migration in neutrophils, integrin-dependent adhesion in T lymphocytes, phagocytosis in macrophages, lysosomes secretion and trafficking at immune synapse in cytolytic effectors and secretory cells, calcium signals and gene transcription in B lymphocytes, natural killer cells and mast cells. The coordination of these different aspects relies on the spatio-temporal organisation of distinct PIP2 pools, generated by the main PIP2 generating enzyme, phosphatidylinositol 4-phosphate 5-kinase (PIP5K). Three different isoforms of PIP5K, named α, β and γ, and different splice variants have been described in leukocyte populations. The isoform-specific coupling of specific isoforms of PIP5K to different families of activating receptors, including integrins, Fc receptors, toll-like receptors and chemokine receptors, is starting to be reported. Furthermore, PIP2 is turned over by multiple metabolising enzymes including phospholipase C (PLC) γ and phosphatidylinositol 3-kinase (PI3K) which, along with Rho family small G proteins, is widely involved in strategic functions within the immune system. The interplay between PIP2, lipid-modifying enzymes and small G protein-regulated signals is also discussed.

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Phosphatidylinositol 4-Phosphate 5-Kinase α Facilitates Toll-like Receptor 4-mediated Microglial Inflammation through Regulation of the Toll/Interleukin-1 Receptor Domain-containing Adaptor Protein (TIRAP) Location*

Cited by 43 publications

References 63 publications

LPS-induced clustering of CD14 triggers generation of PI(4,5)P2

LPS-induced clustering of CD14 triggers generation of PI(4,5)P2

Membrane Targeting of TIRAP is Negatively Regulated by Phosphorylation in its Phosphoinositide-Binding Motif

The multifaceted role of PIP2 in leukocyte biology

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