Plant lipid transfer proteins' ligand enhances allergic sensitization to non-related proteins in murine models

4 Pág.This research was supported by the Spanish Ministry of Science and Innovation through the project LISENTRA, granted by the Spanish Research State Agency (PID2020-113629RB00/AEI/10.13039/501100011033) and FIS grant PI21/00158 (ISCIII). DP-C was granted by Universidad Politécnica de Madrid and Banco Santander for a predoctoral Programa Propio grant. SFB was granted from the ISCIII (FI22/00046). GVP was granted by PRE2021-100446 funded by MCIN/AEI/10.13039/501100011033 and by ESF+. ZG-K and JT-A were granted by funding from the Community of Madrid in the framework of the FOODAL project (S2018/BAA-4574).With funding from the Spanish government through the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2020‐000999‐S)Peer reviewe

Corrigendum: Suitability of potyviral recombinant virus-like particles bearing a complete food allergen for immunotherapy vaccines(Front. Immunol., (2023), 13, (986823), 10.3389/fimmu.2022.986823)

In the published article, there was an error in the author list, and author Marina Amores-Borge was erroneously excluded. The corrected author list appears below. “Diego Pazos-Castro, Clémence Margain, Zulema Gonzalez-Klein, Marina Amores- Borge, Carmen Yuste-Calvo, Maria Garrido-Arandia, Lucia Zurita, Vanesa Esteban, Jaime Tome-Amat, Araceli Diaz-Perales, Fernando Ponz”In the published article, there was an error. The missing author was not included in the Author Contributions section. A correction has been made to Author contributions. This sentence previously stated: “DP-C: Conceptualization, investigation, writing original draft. CM: Investigation. ZG-K: Conceptualization, investigation, writing review. CY-C: Investigation. MG-A: Investigation, writing review. LZ: Investigation. VE: Writing review, resources, funding acquisition. JT-A: Conceptualization, investigation, writing original draft. AD-P: Conceptualization, writing original draft, resources, funding acquisition. FP: Conceptualization, writing review,resources, funding acquisition. All authors contributed to the article and approved the submitted version.” The corrected sentence appears below: “DP-C: Conceptualization, investigation, writing original draft. CM: Investigation. ZG-K: Conceptualization, investigation, writing review. MA-B: Investigation. CY-C: Investigation. MG-A: Investigation, writing review. LZ: Investigation. VE: Writing review, resources, funding acquisition. JT-A: Conceptualization, investigation, writing original draft. AD-P: Conceptualization, writing original draft, resources, funding acquisition. FP: Conceptualization, writing review, resources, funding acquisition. All authors contributed to the article and approved the submitted version.” The authors apologize for these errors and state that they do not change the scientific conclusions of the article in any way. The original article has been updated.Peer reviewe

Identification and molecular characterization of a novel non-specific lipid transfer protein (TdLTP2) from durum wheat

17 Pág.Non-specific lipid transfer proteins (nsLTPs) are small, cysteine-rich proteins, a part of the pathogenesis-related protein family, and numerous of them act as positive regulators during plant disease resistance, growth, and reproduction. These proteins are involved also in the intracellular transfer of lipids, as well as in plant immune responses. Besides their differences in sequences, they show similar features in their structure. However, they show distinct lipid-binding specificities signifying their various biological roles that dictate further structural study. This study reports the identification, in silico characterization and purification of a novel member of the nsLTP2 protein family from durum wheat, TdLTP2. It was generated and purified using the combination of gel filtration chromatography and reverse-phase high-performance liquid chromatography (RP-HPLC). Its identity was detected by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and mass spectrometry (MALDI-TOF). TdLTP2 had been expressed in different stress to detect its localization; therefore, fluor-immunolocalization studies accomplished this data. In this approach, to assess the allergenicity of TdLTP2, thirty patients with baker's asthma were enrolled and ELISA to detect the presence of specific IgE antibodies tested their sera. Moreover, the lipid-binding properties of TdLTP2 were examined in vitro and validated using a molecular docking study. In summary, our results demonstrate a new addition of member in plant nsLTPs family, TdLTP2, which can develop a better understanding about its biological functions and shed light on future applications.This work was funded by the Spanish Ministry of Science and Innovation through the project LISENTRA, granted by the Spanish Research State Agency (PID2020-113629RBI00/AEI/10.13039/501100011033), the Comunidad de Madrid (FOODAL-CM: S2018/BAA4574) Spain, and a grant from the Ministry of Higher Education, and Scientific Research of Tunisia, and a grant from. There was no additional external funding received for this studyPeer reviewe

Comprehensive lipidome of human plasma using minimal sample manipulation by liquid chromatography coupled with mass spectrometry

Comprehensive analysis of the lipidome is addressed here by analysing lipid subclasses not easily detected by current high-throughput methods. Abundant lipid subclasses in human plasma are chromatographically separated from low abundance lipids prior to detection, avoiding the need for derivatisation. Lipid subclasses from the de novo lipogenesis and sphingolipids pathways are presented in this work. Three chromatographic methods here were implemented using a tertiary pumping system to allow for the inclusion of a gradient for analyte separation using A and B pumps, while an isocratic wash elutes interfering compounds. The isocratic wash enabled elution of lipid subclasses not targeted within the method that would otherwise cause background signal in the subsequent sample injection and reduction in column lifetime. Four chromatographic methods coupled with mass spectrometry using targeted and untargeted approaches to separate high and low abundance lipid subclasses are described here. An optimised method for the extraction of lysolipids is also used in addition to Folch extraction in human plasma

Comprehensive lipidome of human plasma using minimal sample manipulation by liquid chromatography coupled with mass spectrometry

12 Pág. Centro de Biotecnología y Genómica de Plantas (CBGP)[Rationale],The present work shows comprehensive chromatographic methods and MS conditions that have been developed based on the chemical properties of each lipid subclass to detect low-abundance molecular species. This study shows that the developed methods can detect low- and/or very-low-abundant lipids like phosphatidic acid (PA) in the glycerophospholipid (GP) method; dihydroceramide (dhCer) and dihydrosphingosine/sphinganine (dhSPB) in the sphingolipid (SP) method; and lysophosphatidic acid (LPA), LPI, LPG and sphingosine-1-phosphate (SPBP) in the lysolipid method.[Methods], An optimised method for the extraction of lysolipids in plasma is used in addition to Folch extraction. Then, four chromatographic methods coupled with mass spectrometry using targeted and untargeted approaches are described here. Three of the methods use a tertiary pumping system to enable the inclusion of a gradient for analyte separation (pumps A and B) and an isocratic wash (pump C). This wash solution elutes interfering compounds that could cause background signal in the subsequent injections, reducing column lifetime.[Results], Semi-quantitative values for 37 lipid subclasses are reported for a plasma sample (NIST SRM 1950). Furthermore, the methods presented here enabled the identification of 338 different lipid molecular species for GPs (mono- and diacyl-phospholipds), SPs, sterols and glycerolipids. The methods have been validated, and the reproducibility is presented here.[Conclusions], The comprehensive analysis of the lipidome addressed here of glycerolipids, GPs, sterols and SPs is in good agreement with previously reported results, in the NIST SRM 1950 sample, by other laboratories. Ten lipid subclasses LPS, LPI, alkyl-lysophosphatidic acid/alkenyl-lysophosphatidic acid, alkyl-lysophosphatidylethanolamine/alkenyl-lysophosphatidylethanolamine, dhCer (d18:0), SPB (d18:1), dhSPB (d18:0) and SPBP (d18:2) have been detected using this comprehensive method and are uniquely reported here.The authors thank BBSRC for funding to the Babraham Institute. Z.G.K. was funded by the Community of Madrid (FOODAL-CM_S2018/BAAA-4574) and received a grant from the Universidad Politécnica de Madrid and Banco Santander for a Programa Propio Mobility grant to work at the Babraham Institute. The authors greatly acknowledge the encouragement, mentoring and conversations with Michael Wakelam during the development of these methods. He is an inspiration to them to carry his love for lipids and MS.Peer reviewe

Plant non-specific lipid transfer proteins: An overview

13 Pág.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.KM was supported through funding by the Ministry of Higher Education and Scientific Research of Tunisia (CP_2019–2022), ZGK was funded by the Community of Madrid (FOODAL-CM_S2018/BAAA-4574). DPC was granted by Universidad Politécnica de Madrid and Banco Santander for a predoctoral Programa Propio grant. GHR was funded by European Commission (H2020-NMBP-X-KET-2017-768641 AllerScreening). JTA was funded by Instituto de Salud Carlos III (ISCIII) co-founded by FEDER Thematic Networks and Cooperative Research Centers: ARADYAL (RD16/0006/0003). This work was also funded by the Spanish Ministry of Science and Innovation (LISSENTRA; PID2020-113629RB-I00).Peer reviewe

Alt a 1 Promotes Allergic Asthma In Vivo Through TLR4-Alveolar Macrophages

12 Pág. Centro de Biotecnología y Genómica de Plantas (CBGP)This research was funded by the Spanish Ministry of Science and Innovation through the project LISENTRA, granted by the Spanish Research State Agency (PID2020-113629RB00/AEI/10.13039/501100011033). GH-R was granted by funding from the European Commission through the project AllerScreening, granted within the R&D framework programme Horizon2020 (H2020-NMBP-X- KET-2017-768641). DP-C was granted by Universidad Politécnica de Madrid and Banco Santander for a predoctoral Programa Propio grant. JT-A and ZG-K were granted by funding from the Community of Madrid included in the project FOODAL (FOODAL-CM; S2018/BAA-4574) co-funded by ESF and ERDF R&D projects call Tecnologías 2018. This work was also supported by Instituto de Salud Carlos III (ISCIII) co-funded by FEDER Thematic Networks and Cooperative Research Centers: ARADYAL (RD16/0006/0003; RD16/0006/0013).Peer reviewe

NLRP3 priming due to skin damage precedes LTP allergic sensitization in a mouse model

Allergic sensitization is initiated by protein and epithelia interaction, although the molecular mechanisms leading this encounter toward an allergic phenotype remain unknown. Here, we apply the two-hit hypothesis of inflammatory diseases to the study of food allergy sensitization. First, we studied the effects of long-term depilation in mice by analyzing samples at different time points. Several weeks of depilation were needed until clear immunological changes were evidenced, starting with upregulation of NLRP3 protein levels, which was followed by overexpression of Il1b and Il18 transcripts. Secondly, we assessed the effects of allergen addition (in this case, Pru p 3 in complex with its natural lipid ligand) over depilated skin. Systemic sensitization was evaluated by intraperitoneal provocation with Pru p 3 and measure of body temperature. Anaphylaxis was achieved, but only in mice sensitized with Prup3_complex and not treated with the NLRP3 inhibitor MCC950, thus demonstrating the importance of both hits (depilation + allergen addition) in the consecution of the allergic phenotype. In addition, allergen encounter (but not depilation) promoted skin remodeling, as well as CD45+ infiltration not only in the sensitized area (the skin), but across several mucosal tissues (skin, lungs, and gut), furtherly validating the systemization of the response. Finally, a low-scale study with human ILC2s is reported, where we demonstrate that Prup3_complex can induce their phenotype switch (↑CD86, ↑S1P1) when cultured in vitro, although more data is needed to understand the implications of these changes in food allergy development.This research was funded by the Spanish Government (MINECO, grant BIO2017-84548- R); Instituto de Salud Carlos III (ISCIII) co-founded by FEDER Thematic Networks and Cooperative Research Centers: ARADYAL (RD16/0006/0003; RD16/0006/0013) and the Community of Madrid (FOODAL-CM_S2018/BAAA-4574). DPC was funded by Universidad Politécnica de Madrid and Banco Santander with a predoctoral Programa Propio grant. ARS was funded from the Community of Madrid included in the project FOOD-AL (FOODAL-CM_S2018/BAAA-4574). GHR was funded by European Comission (H2020-NMBP-X-KET-2017-768641—AllerScreening). JTA was funded from Instituto de Salud Carlos III (ISCIII) co-founded by FEDER Thematic Networks and Cooperative Research Centers: ARADYAL (RD16/0006/0003).Peer reviewe

The key to the allergenicity of lipid transfer protein (LTP) ligands: A structural characterization

Plant lipid transfer proteins are a large family that can be found in all land plants. They have a hydrophobic cavity that allows them to harbor lipids and facilitates their traffic between membranes. However, in humans, this plant protein family is responsible for the main food allergies in the Mediterranean area. Nevertheless, not only the protein itself but also its ligand is relevant for allergic sensitization. The main aim of the present work is to analyse the natural ligands carried by four allergenic LTPs (Tri a 14, Art v 3, Par j 2, and Ole e 7), compared with the previously identified ligand of Pru p 3 (CPT-PHS ligand), and clarify their role within the immunological reactions. Results showed that the ligands of the LTPs studied shared a chemical identity, in which the presence of a polar head was essential to the protein-ligand binding. This ligand was transported through a skin cellular model, and phosphorylated phytosphingosine could be detected as result of cell metabolism. Since sphingosine kinase 1 was overexpressed in keratinocytes incubated with the LTP-ligand complex, this enzyme might be responsible for the phosphorylation of the phytosphingosine fraction of the CPT-PHS ligand. This way, phytosphingosine-1-phosphate could be mimicking the role of the human inflammatory mediator sphingosine-1-phosphate, explaining why LTPs are associated with more severe allergic responses. In conclusion, this work contributes to the understanding of the chemical nature and behavior of lipid ligands carried by allergens, which would help to gain insight into their role during allergic sensitization