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C-type lectin receptors (CLRs), which are pattern recognition receptors responsible for triggering innate immune responses, recognize damaged self-components and immunostimulatory lipids from pathogenic bacteria; however, several of their ligands remain unknown. Here, we propose a new analytical platform combining liquid chromatography-high-resolution tandem mass spectrometry with microfractionation capability (LC-FRC-HRMS/MS) and a reporter cell assay for sensitive activity measurements to develop an efficient methodology for searching for lipid ligands of CLR from microbial trace samples (crude cell extracts of approximately 5 mg dry cell/mL). We also developed an in-house lipidomic library containing accurate mass and fragmentation patterns of more than 10,000 lipid molecules predicted in silico for 90 lipid subclasses and 35 acyl side chain fatty acids. Using the developed LC-FRC-HRMS/MS system, the lipid extracts of Helicobacter pylori were separated and fractionated, and HRMS and HRMS/MS spectra were obtained simultaneously. The fractionated lipid extract samples in 96-well plates were thereafter subjected to reporter cell assays using nuclear factor of activated T cells (NFAT)-green fluorescent protein (GFP) reporter cells expressing mouse or human macrophage-inducible C-type lectin (Mincle). A total of 102 lipid molecules from all fractions were annotated using an in-house lipidomic library. Furthermore, a fraction that exhibited significant activity in the NFAT-GFP reporter cell assay contained α-cholesteryl glucoside, a type of glycolipid, which was successfully identified as a lipid ligand molecule for Mincle. Our analytical platform has the potential to be a useful tool for efficient discovery of lipid ligands for immunoreceptors. [ABSTRACT FROM AUTHOR]

Lipid Transfer Proteins (LTPs) have been described as one of the most prevalent and cross-reactive allergen families in the general population. They are widely distributed among the plant kingdom, as well as in different plant organs ranging from pollen to fruits. Thus, they can initiate allergic reactions with very different outcomes, such as asthma and food allergy. Several mouse models have been developed to unravel the mechanisms that lead LTPs to promote such strong sensitization patterns. Interestingly, the union of certain ligands can strengthen the allergenic capacity of LTPs, suggesting that not only is the protein relevant in the sensitization process, but also the ligands that LTPs carry in their cavity. In fact, different LTPs with pro-allergenic capacity have been shown to transport similar ligands, thus positioning lipids in a central role during the first stages of the allergic response. Here, we offer the latest advances in the use of experimental animals to study the topic, remarking differences among them and providing future researchers a tool to choose the most suitable model to achieve their goals. Also, recent results derived from metabolomic studies in humans are included, highlighting how allergic diseases alter the lipidic metabolism toward a pathogenic state in the individual. Altogether, this review offers a comprehensive body of work that sums up the background evidence supporting the role of lipids as modulators of allergic diseases. Studying the role of lipids during allergic sensitization might broaden our understanding of the molecular events leading to tolerance breakdown in the epithelium, thus helping us to understand how allergy is initiated and established in the individuals.

Plant non-specific lipid transfer proteins (nsLTPs) are usually defined as small, basic proteins, with a wide distribution in all orders of higher plants. Structurally, nsLTPs contain a conserved motif of eight cysteines, linked by four disulphide bonds, and a hydrophobic cavity in which the ligand is housed. This structure confers stability and enhances the ability to bind and transport a variety of hydrophobic molecules. Their highly conserved structural resemblance but low sequence identity reflects the wide variety of ligands they can carry, as well as the broad biological functions to which they are linked to, such as membrane stabilization, cell wall organization and signal transduction. In addition, they have also been described as essential in resistance to biotic and abiotic stresses, plant growth and development, seed development, and germination. Hence, there is growing interest in this family of proteins for their critical roles in plant development and for the many unresolved questions that need to be clarified, regarding their subcellular localization, transfer capacity, expression profile, biological function, and evolution. • nsLTPs is a complex, multigene family evolved in all land plants. • nsLTPs are small, cysteine-rich, very stable proteins with high conserved structure. • nsLTPs bind different lipids in vitro and specific ligands in vivo. • nsLTPs are multifunctional proteins that play key roles in plant development. • nsLTPs are an important allergen family with impact in human health. [ABSTRACT FROM AUTHOR]

In plants, lipid trafficking within and inside the cell is carried out by lipid-binding and transfer proteins. Ligands for these proteins are building and signaling lipid molecules, secondary metabolites with different biological activities due to which they perform diverse functions in plants. Many different classes of such lipid-binding and transfer proteins have been found, but the most common and represented in plants are lipid transfer proteins (LTPs), pathogenesis-related class 10 (PR-10) proteins, acyl-CoA-binding proteins (ACBPs), and puroindolines (PINs). A low degree of amino acid sequence homology but similar spatial structures containing an internal hydrophobic cavity are common features of these classes of proteins. In this review, we summarize the latest known data on the features of these protein classes with particular focus on their ability to bind and transfer lipid ligands. We analyzed the structural features of these proteins, the diversity of their possible ligands, the key amino acids participating in ligand binding, the currently known mechanisms of ligand binding and transferring, as well as prospects for possible application.

Previously, we isolated the lentil allergen Len c 3, belonging to the class of lipid transfer proteins, cross-reacting with the major peach allergen Pru p 3 and binding lipid ligands. In this work, the allergenic capacity of Len c 3 and effects of different lipid ligands on the protein stability and IgE-binding capacity were investigated. Impacts of pH and heat treating on ligand binding with Len c 3 were also studied. It was shown that the recombinant Len c 3 (rLen c 3) IgE-binding capacity is sensitive to heating and simulating of gastroduodenal digestion. While being heated or digested, the protein showed a considerably lower capacity to bind specific IgE in sera of allergic patients. The presence of lipid ligands increased the thermostability and resistance of rLen c 3 to digestion, but the level of these effects was dependent upon the ligand’s nature. The anionic lysolipid LPPG showed the most pronounced protective effect which correlated well with experimental data on ligand binding. Thus, the Len c 3 stability and allergenic capacity can be retained in the conditions of food heat cooking and gastroduodenal digestion due to the presence of certain lipid ligands.

The short palate, lung and nasal epithelial clone 1 (SPLUNC1) protein is a member of the palate, lung, and nasal epithelium clone (PLUNC) family, also known as bactericidal/permeability-increasing (BPI) fold-containing protein, family A, member 1 (BPIFA1). SPLUNC1 is an abundant protein in human airways, but its function remains poorly understood. The lipid ligands of SPLUNC1 as well as other PLUNC family members are largely unknown, although some reports provide evidence that lipopolysaccharide (LPS) could be a lipid ligand. Unlike previous hypotheses, we found significant structural differences between SPLUNC1 and BPI. Recombinant SPLUNC1 produced in HEK 293 cells harbored several molecular species of sphingomyelin and phosphatidylcholine as its ligands. Significantly, in vitro lipid-binding studies failed to demonstrate interactions between SPLUNC1 and LPS, lipoteichoic acid, or polymyxin B. Instead, one of the major and most important pulmonary surfactant phospholipids, dipalmitoylphosphatidylcholine (DPPC), bound to SPLUNC1 with high affinity and specificity. We found that SPLUNC1 could be the first protein receptor for DPPC. These discoveries provide insight into the specific determinants governing the interaction between SPLUNC1 and lipids and also shed light on novel functions that SPLUNC1 and other PLUNC family members perform in host defense. [ABSTRACT FROM AUTHOR]

In plants, lipid trafficking within and inside the cell is carried out by lipid-binding and transfer proteins. Ligands for these proteins are building and signaling lipid molecules, secondary metabolites with different biological activities due to which they perform diverse functions in plants. Many different classes of such lipid-binding and transfer proteins have been found, but the most common and represented in plants are lipid transfer proteins (LTPs), pathogenesis-related class 10 (PR-10) proteins, acyl-CoA-binding proteins (ACBPs), and puroindolines (PINs). A low degree of amino acid sequence homology but similar spatial structures containing an internal hydrophobic cavity are common features of these classes of proteins. In this review, we summarize the latest known data on the features of these protein classes with particular focus on their ability to bind and transfer lipid ligands. We analyzed the structural features of these proteins, the diversity of their possible ligands, the key amino acids participating in ligand binding, the currently known mechanisms of ligand binding and transferring, as well as prospects for possible application., Competing Interests: The authors declare no conflict of interest.

In the endoplasmic reticulum (ER), quality control mechanisms distinguish between correctly and incorrectly folded structures to ensure that aberrant proteins are not processed along the secretory pathway. Numerous studies have demonstrated the functional rescue of ER-retained, aberrant proteins by small membrane permeable molecules called pharmacological chaperones. Pharmacological chaperones can bind to misfolded proteins, including G-protein coupled receptors (GPCRs), and promote their correct folding and export from the ER. Recently, common structural features of GPCRs have been uncovered, including the eighth helical domain in the C-terminal tail and conserved residues in the transmembrane domains. However, little is known about the importance of these features in signaling and intracellular trafficking, because receptors deficient in these domains are likely retained in the ER due to misfolding. In this review, we summarize the current knowledge about the requirement of these consensus domains and amino acid residues for the passing through the quality control of the ER. Furthermore, we propose the utilization of membrane permeable ligands for the transport of their cognate, ER-retained GPCRs to the cell surface. The chaperone activity of these ligands allows us to perform functional analyses of the structure-deficient receptors after their trafficking to the cell surface. © 2010 IUBMB IUBMB Life, 62(6): 457–463, 2010 [ABSTRACT FROM AUTHOR]

Abstract: Acylethanolamides are lipid substances widely distributed in the body, generated from a membrane phospholipid precursor, N-acylphosphatidylethanolamine (NAPE). The recent identification of arachidonoyl ethanolamide (anandamide or AEA) as an endogenous cannabinoid ligand has focused attention on acylethanolamides, which has further increased with the subsequent identification of related additional acylethanolamides with signaling function, such as oleoylethanolamide (OEA) and palmitoylethanolamide (PEA). Most of the biological functions of anandamide are mediated by the two G protein-coupled cannabinoid receptors identified to date, CB1 and CB2, with the transient receptor potential vanilloid-1 receptor being an additional potential target. There has been increasing pharmacological evidence for the existence of additional cannabinoid receptors, with the orphan G protein-coupled receptor GPR55 being the most actively scrutinized, and is one of the subjects of this review. The other receptor reviewed here is GPR119, which can recognize OEA and PEA. These two acylethanolamides, although structurally related to anandamide, do not interact with classical cannabinoid receptors. Instead, they have high affinity for the nuclear receptor PPARα, which is believed to mediate many of their biological effects. [Copyright &y& Elsevier]

Lipid Transfer Proteins (LTPs) have been described as one of the most prevalent and cross-reactive allergen families in the general population. They are widely distributed among the plant kingdom, as well as in different plant organs ranging from pollen to fruits. Thus, they can initiate allergic reactions with very different outcomes, such as asthma and food allergy. Several mouse models have been developed to unravel the mechanisms that lead LTPs to promote such strong sensitization patterns. Interestingly, the union of certain ligands can strengthen the allergenic capacity of LTPs, suggesting that not only is the protein relevant in the sensitization process, but also the ligands that LTPs carry in their cavity. In fact, different LTPs with pro-allergenic capacity have been shown to transport similar ligands, thus positioning lipids in a central role during the first stages of the allergic response. Here, we offer the latest advances in the use of experimental animals to study the topic, remarking differences among them and providing future researchers a tool to choose the most suitable model to achieve their goals. Also, recent results derived from metabolomic studies in humans are included, highlighting how allergic diseases alter the lipidic metabolism toward a pathogenic state in the individual. Altogether, this review offers a comprehensive body of work that sums up the background evidence supporting the role of lipids as modulators of allergic diseases. Studying the role of lipids during allergic sensitization might broaden our understanding of the molecular events leading to tolerance breakdown in the epithelium, thus helping us to understand how allergy is initiated and established in the individuals., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Gonzalez-Klein, Pazos-Castro, Hernandez-Ramirez, Garrido-Arandia, Diaz-Perales and Tome-Amat.)

Previously, we isolated the lentil allergen Len c 3, belonging to the class of lipid transfer proteins, cross-reacting with the major peach allergen Pru p 3 and binding lipid ligands. In this work, the allergenic capacity of Len c 3 and effects of different lipid ligands on the protein stability and IgE-binding capacity were investigated. Impacts of pH and heat treating on ligand binding with Len c 3 were also studied. It was shown that the recombinant Len c 3 (rLen c 3) IgE-binding capacity is sensitive to heating and simulating of gastroduodenal digestion. While being heated or digested, the protein showed a considerably lower capacity to bind specific IgE in sera of allergic patients. The presence of lipid ligands increased the thermostability and resistance of rLen c 3 to digestion, but the level of these effects was dependent upon the ligand's nature. The anionic lysolipid LPPG showed the most pronounced protective effect which correlated well with experimental data on ligand binding. Thus, the Len c 3 stability and allergenic capacity can be retained in the conditions of food heat cooking and gastroduodenal digestion due to the presence of certain lipid ligands. [ABSTRACT FROM AUTHOR]

The short palate, lung and nasal epithelial clone 1 (SPLUNC1) protein is a member of the palate, lung, and nasal epithelium clone (PLUNC) family, also known as bactericidal/permeability-increasing (BPI) fold-containing protein, family A, member 1 (BPIFA1). SPLUNC1 is an abundant protein in human airways, but its function remains poorly understood. The lipid ligands of SPLUNC1 as well as other PLUNC family members are largely unknown, although some reports provide evidence that lipopolysaccharide (LPS) could be a lipid ligand. Unlike previous hypotheses, we found significant structural differences between SPLUNC1 and BPI. Recombinant SPLUNC1 produced in HEK 293 cells harbored several molecular species of sphingomyelin and phosphatidylcholine as its ligands. Significantly, in vitro lipid-binding studies failed to demonstrate interactions between SPLUNC1 and LPS, lipoteichoic acid, or polymyxin B. Instead, one of the major and most important pulmonary surfactant phospholipids, dipalmitoylphosphatidylcholine (DPPC), bound to SPLUNC1 with high affinity and specificity. We found that SPLUNC1 could be the first protein receptor for DPPC. These discoveries provide insight into the specific determinants governing the interaction between SPLUNC1 and lipids and also shed light on novel functions that SPLUNC1 and other PLUNC family members perform in host defense., (© FASEB.)