Nonspecific Lipid Transfer Protein Research Articles

IgE-mediated reactivity to non-specific lipid transfer protein (nsLTP): clinical implications and management ‒ a consensus document of the Association of Italian Territorial and Hospital Allergists and Immunologists (AAIITO).

The primary cause of adult-onset food allergy in Mediterranean countries is IgE-mediated reactivity to non-specific Lipid Transfer Protein (nsLTP), with a prevalence of 9.5% in Italy. nsLTP is heat- and pepsin-stable due to its 3D structure, causing severe allergic reactions, even anaphylaxis. It's conserved across plants and a "panallergen" due to homologous forms in various vegetable foods. Found in Rosaceae fruits' skin, it's categorized into nsLTP1 (9 kDa) and nsLTP2 (7 kDa), representing 93% and 7% of the molecules described to date, respectively. Pru p 3 (nsLTP1) from peach is a primary sensitizer, binding more epitopes than other homologs. Cross-reactivity varies in sensitized patients, influenced by IgE levels. Clinical manifestations range from none to various symptoms. Managing patients sensitized to nsLTP without clinical allergy is a challenge. Sensitization hierarchy usually starts with peach, then expands through Prunoideae, Rosaceae, and other foods. Clinical symptoms don't always expand across LTPs. Patients can tolerate some nsLTP-containing foods and consuming them may maintain tolerance. The absence of guidelines led to the Associazione Allergologi Immunologi Italiani Territoriali e Ospedalieri (AAIITO) creating a consensus-based document. Strategies involve avoidance, self-injectable adrenaline, verification through in vivo and in vitro testing, considering cofactors, and peeling fruits. In localized reactions, abstinence is recommended if specific IgE is high. Concurrent pollinosis may complicates diagnosis, but may help management since symptoms are often less severe. Asymptomatic patients are advised to continue normal diets while considering cofactors and total IgE levels. Management strategies should be case-specific, based on expert Consensus Document.

Factors and co-factors influencing clinical manifestations in nsLTPs allergy: between the good and the bad.

Non-specific lipid transfer proteins (nsLTPs) are a family of plant pan-allergens that represent the primary cause of food allergies in the Mediterranean area, characterized by a wide range of clinical manifestations, ranging from the total absence of symptoms up to anaphylaxis. This wide variety of symptoms is related to the intrinsic capacity of nsLTPs to cause an allergic reaction in a specific subject, but also to the presence of co-factors exacerbating (i.e., exercise, NSAIDs, PPIs, alcohol, cannabis, prolonged fasting, menstruation, acute infections, sleep deprivation, chronic urticaria) or protecting from (i.e., co-sensitization to PR10, profilin or polcalcin) severe reactions. In this picture, recognizing some nsLTPs-related peculiarities (i.e., route, type and number of sensitizations, concentration of the allergen, cross-reactions) and eventual co-factors may help the allergist to define the risk profile of the single patient, in order to promote the appropriate management of the allergy from dietary advices up to the prescription of life-saving epinephrine autoinjector.

Identification and evolution of nsLTPs in the root nodule nitrogen fixation clade and molecular response of Frankia to AgLTP24

Non-specific lipid transfer proteins (nsLTPs) are antimicrobial peptides, involved in several plant biological processes including root nodule nitrogen fixation (RNF). Nodulating plants belonging to the RNF clade establish symbiosis with the nitrogen-fixing bacteria rhizobia (legumes symbiosis model) and Frankia (actinorhizal symbiosis model) leading to root nodule formation. nsLTPs are involved in processes active in early step of symbiosis and functional nodule in both models. In legumes, nsLTPs have been shown to regulate symbiont entry, promote root cortex infection, membrane biosynthesis, and improve symbiosis efficiency. More recently, a nsLTP, AgLTP24 has been described in the context of actinorhizal symbiosis between Alnus glutinosa and Frankia alni ACN14a. AgLTP24 is secreted at an early step of symbiosis on the deformed root hairs and targets the symbiont in the nitrogen-fixing vesicles in functional nodules. nsLTPs are involved in RNF, but their functions and evolutionary history are still largely unknown. Numerous putative nsLTPs were found up-regulated in functional nodules compared to non-infected roots in different lineages within the RNF clade. Here, results highlight that nodulating plants that are co-evolving with their nitrogen-fixing symbionts appear to have independently specialized nsLTPs for this interaction, suggesting a possible convergence of function, which opens perspectives to investigate nsLTPs functions in RNF.

Antifungal peptides from faba bean flour fermented by Levilactobacillus brevis AM7 improve the shelf-life of composite faba-wheat bread

Faba bean is an important crop from an ecological, nutritional, and economical perspective, hence its incorporation in plant-based products can play a key role in the sustainability of the whole food system. In this study, the antifungal properties of faba bean flour were investigated employing three bioprocessing treatments including hydrolysis with a commercial proteolytic enzyme preparation, and fermentation with selected or commercial mixed sourdough starters. Faba bean flour fermented by Levilactobacillus brevis AM7 showed the broadest inhibitory spectrum against the fungal species tested and antifungal activity ranging from 25 to 73 %. Bioactive proteins and peptides were found to be responsible for the antifungal activity. Polypeptides sequences corresponding to defensin-like and non-specific lipid-transfer proteins and seven antifungal peptides having 11–22 amino acid residues and mass from 1240.7 to 2624.2 Da, were identified via Liquid Chromatography-Electrospray Ionisation-Mass Spectra/Mass Spectra (nano-LC-ESI-MS/MS). To assess the antifungal effect in food processing conditions, fermented faba bean was used as ingredient in breadmaking at a substitution level of 30 % of the dough weight. Wheat composite-bread containing faba bean sourdough, which could be claimed as “source of proteins”, according to the current EC Regulation, delayed the molds contamination, lasting up to 10 days before mold appearance, and behaving similarly to wheat bread containing calcium propionate.

Proteomics-Based Approach for Detailing the Allergenic Profile of Cannabis Chemotypes.

Allergic sensitization to cannabis is an emerging public health concern and is difficult to clinically establish owing to lack of standardized diagnostic approaches. Attempts to develop diagnostic tools were largely hampered by the Schedule I restrictions on cannabis, which limited accessibility for research. Recently, however, hemp was removed from the classified list, and increased accessibility to hemp allows for the evaluation of its practical clinical value for allergy diagnosis. We hypothesized that the proteomic profile is preserved across different cannabis chemotypes and that hemp would be an ideal source of plant material for clinical testing. Using a proteomics-based approach, we examined whether distinct varieties of cannabis plant contain relevant allergens of cannabis. Cannabis extracts were generated from high tetrahydrocannabinol variety (Mx), high cannabidiol variety (V1-19) and mixed profile variety (B5) using a Plant Total Protein Extraction Kit. Hemp extracts were generated using other standardized methods. Protein samples were subjected to nanoscale tandem mass spectrometry. Acquired peptides sequences were examined against the Cannabis sativa database to establish protein identity. Non-specific lipid transfer protein (Can s 3) level was measured using a recently developed ELISA 2.0 assay. Proteomic analysis identified 49 distinct potential allergens in protein extracts from all chemotypes. Most importantly, clinically relevant and validated allergens, such as profilin (Can s 2), Can s 3 and Bet v 1-domain-containing protein 10 (Can s 5), were identified in all chemotypes at label-free quantification (LFP) intensities > 106. However, the oxygen evolving enhancer protein 2 (Can s 4) was not detected in any of the protein samples. Similarly, Can s 2, Can s 3 and Can s 5 peptides were also detected in hemp protein extracts. The validation of these findings using the ELISA 2.0 assay indicated that hemp extract contains 30-37 ng of Can s 3 allergen per µg of total protein. Our proteomic studies indicate that relevant cannabis allergens are consistently expressed across distinct cannabis chemotypes. Further, hemp may serve as an ideal practical substitute for clinical testing, since it expresses most allergens relevant to cannabis sensitization, including the validated major allergen Can s 3.

Potato Non-Specific Lipid Transfer Protein StnsLTPI.33 Is Associated with the Production of Reactive Oxygen Species, Plant Growth, and Susceptibility to Alternaria solani.

Plant non-specific lipid transfer proteins (nsLTPs) are small proteins capable of transferring phospholipids between membranes and binding non-specifically fatty acids in vitro. They constitute large gene families in plants, e.g., 83 in potato (Solanum tuberosum). Despite their recognition decades ago, very few have been functionally characterized. Here, we set out to better understand the function of one of the potato members, StnsLTPI.33. Using quantitative polymerase chain reaction, we show that StnsLTPI.33 is expressed throughout the potato plant, but at relatively higher levels in roots and leaves compared to petals, anthers, and the ovary. We also show that ectopically-expressed StnsLTPI.33 fused to green fluorescent protein colocalized with an apoplastic marker in Nicotiana benthamiana leaves, indicating that StnsLTPI.33 is targeted to the apoplast. Constitutive overexpression of the StnsLTPI.33 gene in potato led to increased levels of superoxide anions and reduced plant growth, particularly under salt stress conditions, and enhanced susceptibility to Alternaria solani. In addition, StnsLTPI.33-overexpressing plants had a depleted leaf pool of pipecolic acid, threonic acid, and glycine, while they accumulated putrescine. To our knowledge, this is the first report of an nsLTP that is associated with enhanced susceptibility to a pathogen in potato.

Lipid transfer protein, OsLTPL18, is essential for grain weight and seed germination in rice

Nonspecific lipid transfer proteins (nsLTPs) promote the intermembrane transportation of phospholipids, fatty acids, and steroids, and play diverse roles in various biological processes. However, the potential roles of the rice nsLTPs have not been well elucidated yet. Here, the functions of OsLTPL18 were analyzed using CRISPR/Cas9 strategy and cytological analysis. The osltpl18 (osltpl18-1, osltpl18-2, and osltpl18-3) seeds were thinner, and 1000-grain weight and grain thickness of osltpl18 plants were decreased obviously, compared to the ZH11. Meanwhile, the results of germination assay and 1 % TTC staining showed that vigor of osltpl18 seeds decreased significantly. Furthermore, the results of scanning electron microscopy (SEM) revealed that the cell width of spikelet hull in osltpl18 lines was significantly reduced than that in WT, as well as cell number in grain-width direction. Finally, we found that co-expressed genes were enriched in glucan biosynthesis, protein transporter activity, serine-type endopeptidase inhibitor activity, and nutrient reservoir activity. In this study, we discussed that OsLTPL18 might have coordinating functions in regulation of grain weight and germination in rice.

The structural characterization and bioactivity assessment of nonspecific lipid transfer protein 1 (nsLTP1) from caraway (Carum carvi) seeds

BackgroundCarum carvi (caraway) of the Apiaceae family has been used in many cultures as a cooking spice and part of the folk medicine. Previous reports primarily focus on the medicinal properties of caraway seed essential oil and the whole seeds extract. However, no effort has been made to study caraway proteins and their potential pharmacological properties, including nonspecific lipid transfer protein (nsLTP), necessitating further research. The current study aimed to characterize nonspecific lipid transfer protein 1 (nsLTP1) from caraway seed, determine its three-dimensional structure, and analyze protein–ligand complex interactions through docking studies. We also evaluated nsLTP1 in vitro cytotoxic effect and antioxidant capacity. Additionally, nsLTP1 thermal- and pH- stability were investigated.MethodsCaraway nsLTP1 was purified using two-dimensional chromatography. The complete amino acid sequence of nsLTP1 was achieved by intact protein sequence for the first 20 residues and the overlapping digested peptides. The three-dimensional structure was predicted using MODELLER. Autodock Vina software was employed for docking fatty acids against caraway nsLTP1. Assessment of nsLTP1 cytotoxic activity was achieved by MTS assay, and the Trolox equivalent antioxidant capacity (TAC) was determined. Thermal and pH stability of the nsLTP1 was examined by circular dichroism (CD) spectroscopy.ResultsCaraway nsLTP1 is composed of 91 residues and weighs 9652 Da. The three-dimensional structure of caraway nsLTP1 sequence was constructed based on searching known structures in the PDB. We chose nsLTP of Solanum melongena (PDB ID: 5TVI) as the modeling template with the highest identity among all other homologous proteins. Docking linolenic acid with caraway protein showed a maximum binding score of -3.6 kcal/mol. A preliminary screening of caraway nsLTP1 suppressed the proliferation of human breast cancer cell lines MDA-MB-231 and MCF-7 in a dose‑dependent manner with an IC50 value of 52.93 and 44.76 μM, respectively. Also, nsLTP1 (41.4 μM) showed TAC up to 750.4 μM Trolox equivalent. Assessment of nsLTP1 demonstrated high thermal/pH stability.ConclusionTo the best of our knowledge, this is the first study carried out on nsLTP1 from caraway seeds. We hereby report the sequence of nsLTP1 from caraway seeds and its possible interaction with respective fatty acids using in silico approach. Our data indicated that the protein had anticancer and antioxidant activities and was thermally stable.

An Overview of Fruit Allergens: Structural, Functional, Phylogenetical, and Clinical Aspects

Most of the allergenic proteins from fruits identified so far belong to different families of pathogenesis-related (PR) proteins. These PR proteins have been classified in different families of structurally and functionally unrelated proteins, but the majority of all fruit allergens belong to three groups, in particular PR-5 thaumatin-like proteins (TLP), PR-10 Bet v 1-like proteins, and PR-14 non-specific lipid transfer proteins (nsTLP). Some allergenic proteins from fruits can also be found among PR-protein families of PR-2 β1,3-glucanase proteins, PR-3 chitinases I, II, IV–VII, and PR-8 chitinases III. In addition, other important fruit allergens occur in protein families unrelated to the PR-protein families, such as the profilins and the newly emerging group of gibberellin-regulated proteins (GBRP). Finally, proteins that belong to seed storage proteins from higher plants, including 2S albumins, 7S globulins (vicilin), and 11S globulins (legumin), must be retained as possible potential fruit allergens resulting from the unintended consumption of the seeds. Here, we present an overview of the structural organization, functional properties, and phylogenetical relationships among these different groups of fruit allergens, supporting the occurrence of cross-reactivity and cross-allergenicity often described between fruit allergens, and the corresponding allergens from vegetables and pollens.

Comparative Transcriptome Analysis Reveals Novel Candidate Resistance Genes Involved in Defence against Phytophthora cactorum in Strawberry.

Crown rot, caused by Phytophthora cactorum, is a devastating disease of strawberry. While most commercial octoploid strawberry cultivars (Fragaria × ananassa Duch) are generally susceptible, the diploid species Fragaria vesca is a potential source of resistance genes to P. cactorum. We previously reported several F. vesca genotypes with varying degrees of resistance to P. cactorum. To gain insights into the strawberry defence mechanisms, comparative transcriptome profiles of two resistant genotypes (NCGR1603 and Bukammen) and a susceptible genotype (NCGR1218) of F. vesca were analysed by RNA-Seq after wounding and subsequent inoculation with P. cactorum. Differential gene expression analysis identified several defence-related genes that are highly expressed in the resistant genotypes relative to the susceptible genotype in response to P. cactorum after wounding. These included putative disease resistance (R) genes encoding receptor-like proteins, receptor-like kinases, nucleotide-binding sites, leucine-rich repeat proteins, RPW8-type disease resistance proteins, and 'pathogenesis-related protein 1'. Seven of these R-genes were expressed only in the resistant genotypes and not in the susceptible genotype, and these appeared to be present only in the genomes of the resistant genotypes, as confirmed by PCR analysis. We previously reported a single major gene locus RPc-1 (Resistance to Phytophthora cactorum 1) in F. vesca that contributed resistance to P. cactorum. Here, we report that 4-5% of the genes (35-38 of ca 800 genes) in the RPc-1 locus are differentially expressed in the resistant genotypes compared to the susceptible genotype after inoculation with P. cactorum. In particular, we identified three defence-related genes encoding wall-associated receptor-like kinase 3, receptor-like protein 12, and non-specific lipid-transfer protein 1-like that were highly expressed in the resistant genotypes compared to the susceptible one. The present study reports several novel candidate disease resistance genes that warrant further investigation for their role in plant defence against P. cactorum.

Improving In Vitro Detection of Sensitization to Lipid Transfer Proteins: A New Molecular Multiplex IgE Assay.

LTP-syndrome is characterized by sensitization (IgE) to multiple non-specific lipid transfer proteins (nsLTPs) with a variable clinical outcome. The treatment is primarily based on offending food avoidance. However, the determination of Pru p 3-specific IgE is currently the main diagnostic tool to assess sensitization to nsLTPs. Herein, the study evaluates improvement of LTP-syndrome diagnosis and clinical management using a new IgE multiplex-immunoblot assay with a high diversity of food nsLTPs. An EUROLINE-LTP strip with 28 recombinant nsLTPs from 18 allergenic sources is designed. In total the study investigates 38 patients with LTP-syndrome and compares results from the nsLTPs (LTP-strip) with the respective food extracts of Prick-by-prick (PbP) testing. The agreement exceeds 70% for most nsLTPs, e.g., Pru p 3 (100%), Mal d 3 (97%), Pru av 3 (89%), Pha v 3 isoforms (87%/84%), Ara h 9 (82%), Cor a 8 (82%), and Jug r 3 (82%). The functionality and allergenic relevance of nine recombinant nsLTPs are proven by Basophil activation testing (BAT). The new IgE multiplex-immunoblot nsLTP assay shows a good diagnostic performance allowing culprit food assessment. Negative results from LTP-strip may indicate potentially tolerable foods, improving diet intervention and patients' quality of life.

Sequence Analysis and Biochemical Characteristics of Two Non-specific Lipid Transfer Proteins from Tartary Buckwheat Seeds.

Plant non-specific lipid transfer proteins (nsLTPs) play an important role in plant resistance to various stresses, and show potential applications in agriculture, industrial manufacturing, and medicine. In addition, as more and more nsLTPs are identified as allergens, nsLTPs have attracted interest due to their allergenicity. Two nsLTPs from Tartary buckwheat have been isolated and identified. There is a need to study their biochemical characteristics and allergenicity. The study aims to investigate the biochemical characteristics of two nsLTPs from Tartary buckwheat seeds and evaluate their potential allergenicity. Two nsLTPs derived from Tartary buckwheat, namely FtLTP1a and FtLTP1b, were produced by gene cloning, expression, and purification. Sequence analysis and biochemical characteristics of the proteins, including lipid binding ability, α-amylase inhibition activity, antifungal activity, and allergenic activity, were investigated. High-purity recombinant FtLTP1a and FtLTP1b were obtained. FtLTP1a and FtLTP1b exhibited similar lipid binding and antifungal properties. Only FtLTP1b showed weak inhibitory activity against α-amylase. FtLTP1b could specifically bind IgE in the serum allergic to buckwheat and cross-react with pollen (w6). FtLTP1b is a novel allergenic member of the lipid-transfer protein 1 family found in Tartary buckwheat.

The non-specific lipid transfer protein McLTPII.9 of Mentha canadensis is involved in peltate glandular trichome density and volatile compound metabolism.

Mentha canadensis L. is an important spice crop and medicinal herb with high economic value. The plant is covered with peltate glandular trichomes, which are responsible for the biosynthesis and secretion of volatile oils. Plant non-specific lipid transfer proteins (nsLTPs) belong to a complex multigenic family involved in various plant physiological processes. Here, we cloned and identified a non-specific lipid transfer protein gene (McLTPII.9) from M. canadensis, which may positively regulate peltate glandular trichome density and monoterpene metabolism. McLTPII.9 was expressed in most M. canadensis tissues. The GUS signal driven by the McLTPII.9 promoter in transgenic Nicotiana tabacum was observed in stems, leaves, and roots; it was also expressed in trichomes. McLTPII.9 was associated with the plasma membrane. Overexpression of McLTPII.9 in peppermint (Mentha piperita. L) significantly increased the peltate glandular trichome density and total volatile compound content compared with wild-type peppermint; it also altered the volatile oil composition. In McLTPII.9-overexpressing (OE) peppermint, the expression levels of several monoterpenoid synthase genes and glandular trichome development-related transcription factors-such as limonene synthase (LS), limonene-3-hydroxylase (L3OH), geranyl diphosphate synthase (GPPS), HD-ZIP3, and MIXTA-exhibited varying degrees of alteration. McLTPII.9 overexpression resulted in both a change in expression of genes for terpenoid biosynthetic pathways which corresponded with an altered terpenoid profile in OE plants. In addition, peltate glandular trichome density was altered in the OE plants as well as the expression of genes for transcription factors that were shown to be involved in trichome development in plants.

Non-specific LIPID TRANSFER PROTEIN 1 enhances immunity against tobacco mosaic virus in Nicotiana benthamiana.

Plant non-specific lipid transfer proteins (nsLTPs) are small, cysteine-rich proteins that play significant roles in biotic and abiotic stress responses; however, the molecular mechanism of their functions against viral infections remains unclear. In this study, we employed virus-induced gene-silencing and transgenic overexpression to functionally analyse a type-I nsLTP in Nicotiana benthamiana, NbLTP1, in the immunity response against tobacco mosaic virus (TMV). NbLTP1 was inducible by TMV infection, and its silencing increased TMV-induced oxidative damage and the production of reactive oxygen species (ROS), compromised local and systemic resistance to TMV, and inactivated the biosynthesis of salicylic acid (SA) and its downstream signaling pathway. The effects of NbLTP1-silencing were partially restored by application of exogenous SA. Overexpressing NbLTP1 activated genes related to ROS scavenging to increase cell membrane stability and maintain redox homeostasis, confirming that an early ROS burst followed by ROS suppression at the later phases of pathogenesis is essential for resistance to TMV infection. The cell-wall localization of NbLTP1 was beneficial to viral resistance. Overall, our results showed that NbLTP1 positively regulates plant immunity against viral infection through up-regulating SA biosynthesis and its downstream signaling component, NONEXPRESSOR OF PATHOGENESIS-RELATED 1 (NPR1), which in turn activates pathogenesis-related genes, and by suppressing ROS accumulation at the later phases of viral pathogenesis.

A non-specific lipid transfer protein, NtLTPI.38, positively mediates heat tolerance by regulating photosynthetic ability and antioxidant capacity in tobacco

Non-specific lipid transfer proteins (nsLTPs) play an important role in plant growth and stress resistance; however, their function in tobacco remains poorly understood. Therefore, to explore the function of NtLTP in response to high temperature, we identified an NtLTPI.38 from tobacco, obtained its overexpression and knockout transgenic plants, and further studied their response to heat stress (42 °C). The results showed that NtLTPI.38 overexpression in tobacco reduced chlorophyll degradation, alleviated the high temperature damage to photosynthetic organs, and enhanced the photosynthetic capacity of tobacco under heat stress. NtLTPI.38 overexpression in heat-stressed tobacco increased the contents of soluble sugar and protein, proline, and flavonoid substances, reduced the relative conductivity, and decreased H2O2, O2•−, and MDA accumulation, and increased the enzymatic antioxidant activities, such as superoxide dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX), compared to wild type (WT) and knockout mutant plants. RT-PCR confirmed that the expression levels of antioxidant enzymes and thermal stress-related genes were significantly upregulated under thermal stress in overexpression plants. Therefore, NtLTPI.38 enhanced heat tolerance in tobacco by mitigating photosynthetic damage and improving osmoregulation and antioxidant capacity. These results provided the theoretical basis and a potential resource for further breeding projects to improve heat tolerance in plants.

From Gene to Transcript and Peptide: A Deep Overview on Non-Specific Lipid Transfer Proteins (nsLTPs).

Non-specific lipid transfer proteins (nsLTPs) stand out among plant-specific peptide superfamilies due to their multifaceted roles in plant molecular physiology and development, including their protective functions against pathogens. These antimicrobial agents have demonstrated remarkable efficacy against bacterial and fungal pathogens. The discovery of plant-originated, cysteine-rich antimicrobial peptides such as nsLTPs has paved the way for exploring the mentioned organisms as potential biofactories for synthesizing antimicrobial compounds. Recently, nsLTPs have been the focus of a plethora of research and reviews, providing a functional overview of their potential activity. The present work compiles relevant information on nsLTP omics and evolution, and it adds meta-analysis of nsLTPs, including: (1) genome-wide mining in 12 plant genomes not studied before; (2) latest common ancestor analysis (LCA) and expansion mechanisms; (3) structural proteomics, scrutinizing nsLTPs' three-dimensional structure/physicochemical characteristics in the context of nsLTP classification; and (4) broad nsLTP spatiotemporal transcriptional analysis using soybean as a study case. Combining a critical review with original results, we aim to integrate high-quality information in a single source to clarify unexplored aspects of this important gene/peptide family.

NtLTPI.38, a plasma membrane-localized protein, mediates lipid metabolism and salt tolerance in Nicotiana tabacum

Non-specific lipid transfer proteins (nsLTPs) typically have conserved structural resemblance, low sequence identity, and broad biological functions in plant growth and stress resistance. Here, a plasma membrane-localized nsLTP, NtLTPI.38, was identified in tobacco plants. Multi-omics integrated analysis revealed that NtLTPI.38 overexpression or knock out significantly changed glycerophospholipid and glycerolipid metabolism pathways. NtLTPI.38 overexpression remarkably increased phosphatidylcholine, phosphatidylethanolamine, triacylglycerol, and flavonoid levels, but decreased ceramides compared to wild type and mutant lines. Differentially expressed genes were associated with lipid metabolite and flavonoid synthesis. Many genes related to Ca2+ channels, abscisic acid (ABA) signal transduction, and ion transport pathways were upregulated in overexpressing plants. NtLTPI.38 overexpression in salt-stressed tobacco triggered a Ca2+ and K+ influx in leaves, increased the contents of chlorophyll, proline, flavonoids, and osmotic tolerance, and raised enzymatic antioxidant activities as well as the expression level of related genes. However, mutants accumulated more O2− and H2O2, exhibited ionic imbalance, gathered excess Na+, Cl−, and malondialdehyde, with more severe ion leakage. Therefore, NtLTPI.38 enhanced salt tolerance in tobacco by regulating lipid and flavonoid synthesis, antioxidant activity, ion homeostasis, and ABA signaling pathways.

Lysine malonylation of DgnsLIPID TRANSFER PROTEIN1 at the K81 site improves cold resistance in chrysanthemum.

Lysine malonylation (Kmal) is a recently discovered posttranslational modification, and its role in the response to abiotic stress has not been reported in plants. In this study, we isolated a nonspecific lipid transfer protein, DgnsLTP1, from chrysanthemum (Dendranthema grandiflorum var. Jinba). Overexpression and CRISPR-Cas9-mediated gene editing of DgnsLTP1 demonstrated that the protein endows chrysanthemum with cold tolerance. Yeast 2-hybrid, bimolecular fluorescence complementation, luciferase complementation imaging, and coimmunoprecipitation experimental results showed that DgnsLTP1 interacts with a plasma membrane intrinsic protein (PIP) DgPIP. Overexpressing DgPIP boosted the expression of DgGPX (glutathione peroxidase), increased the activity of GPX, and decreased the accumulation of reactive oxygen species (ROS), thereby enhancing the low-temperature stress tolerance of chrysanthemum, while the CRISPR-Cas9-mediated mutant dgpip inhibited this process. Transgenic analyses in chrysanthemum showed that DgnsLTP1 improves the cold resistance of chrysanthemum in a DgPIP-dependent manner. Moreover, Kmal of DgnsLTP1 at the K81 site prevented the degradation of DgPIP in Nicotiana benthamiana and chrysanthemum, further promoted DgGPX expression, enhanced GPX activity, and scavenged excess ROS produced by cold stress, thereby further enhancing the cold resistance of chrysanthemum.

Sensitization to nsLTP: A Retrospective Study in An Italian Pediatric Population over the Last Decade.

Food allergy is common in the Mediterranean, especially concerning lipid transfer proteins (LTPs) allergy. LTPs are widespread plant food allergens in fruits, vegetables, nuts, pollen, and latex. Also, LTPs are prevalent food allergens in the Mediterranean area. They can sensitize via the gastrointestinal tract and cause a wide range of conditions: from mild reactions, such as oral allergy syndrome, to severe reactions, such as anaphylaxis. LTP allergy in the adult population is well described in the literature, concerning both the prevalence and clinical characteristics. However, there is poor knowledge about its prevalence and clinical manifestation in children living in the Mediterranean. This study, including 800 children aged from 1 to 18 years, investigated the prevalence of 8 different molecules of nonspecific LTP over time in an Italian pediatric population visited over the last 11 years. About 52% of the test population was sensitized to at least one LTP molecule. For all the LTPs analyzed, sensitization increased over time. In particular, using the years 2010 through 2020 as a comparison, the major increases were observed for the LTPs of the English walnut Jug r 3, the peanut Ara h 9, and the plane tree Pla a 3 (about 50%); the increase of the LTP of the Hazelnut Cor a 8 was about 36%, and that of the LTP of the artemisia Art v 3 was approximately 30%. The latest evidence in the literature indicates an increase in food allergy prevalence in the general population, including children. Therefore, the present survey represents an interesting perspective about the pediatric population of the Mediterranean area, exploring the trend of LTP allergy.

Identification of nsLTP family in Chinese white pear (Pyrus bretschneideri) reveals its potential roles in russet skin formation

Identification of PbLTP genes in pear and functional characterization of PbLTP4 in the transport of suberin monomers of russet skin formation. Non-specific lipid-transfer protein (nsLTP) is an abundant and diverse alkaline small molecule protein in the plant kingdom with complex and diverse biophysiological functions, such as transfer of phospholipids, reproductive development, pathogen defence and abiotic stress response. Up to now, only a tiny fraction of nsLTPs have been functionally identified, and the distribution of nsLTPs in pear (Pyrus bretschneideri) (PbLTPs) has not been fully characterized. In this study, the genome-wide analysis of the nsLTP gene family in the pear genome identified 67 PbLTP proteins, which could be divided into six types (1, 2, C, D, E, and G). Similar intron/exon structural patterns were observed in the same type, strongly supporting their close evolutionary relationship. In addition, PbLTP4 was highly expressed in russet pear skin compared with green skin, which was located in the plasma membrane. Coexpression network analysis showed that PbLTP4 closely related to suberin biosynthetic genes. The biological function of PbLTP4 in promoting suberification has been demonstrated by overexpression in Arabidopsis. Identification of suberin monomers showed that PbLTP4 promotes suberification by regulating 9,12-octadecadienoic acid and hexadecanoic acid transport. These results provide helpful insights into the characteristics of PbLTP genes and their biological function in the transport of suberin monomers of russet skin formation.