Oleaceae Molecular Allergens Mayte Villalba, RFAM-4, Paris 2018 Which Factors Affect a Patient with Pollinosis?
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Oleaceae molecular allergens Mayte Villalba, RFAM-4, Paris 2018 Which factors affect a patient with pollinosis? POLLEN Pollen type Concentration Season Transport ALLERGEN ENVIRONMENT Structure Other allergens Function Cofactors Stability Contaminants Tobacco smoke PATIENT Genetics Epigenetics Altered Epithelium pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Plant species with allergenic pollens in Spain Allergenic pollens: Cupressaceae, Platanaceae, Poaceae, Chenopodiaceae, Oleaceae 1. The overlap of Olive Privet Ash Lilac pollination periods of Cypress Forsythia several species Platanus 3. Strong pollen gradient from Grasses nord to south of Spain Olive Chenopodiaceae 2. Different pollination intervals between geographical areas Barber et al. Curr Allergy Asthma Rep 2015 pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Oleaceae family • Family of flowering plants in the order Lamiales • Contains 25 genera and more than 700 species • Trees, shrubs, hedges • Grow spontaneously (Olea oleaster) or can be cultivated • Ornamental, medical, food purposes • Adaptation to different grounds http://www.mobot.org/mobot/research pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Which Oleaceae species are allergenic? Forsythia Ash Privet Olive Lilac Forsythia Fraxinus Ligustrum Olea Syringa intermedia excelsior vulgare europaea vulgaris Pollen sensitization is usually restricted to anemophilous plants, mainly Ash and Olive. Anemophilous pollens are small, dehydrated with good aerodynamic properties that allow their dissemination over hundreds of kilometers. Privet, lilac, and forsythia which are entomophilous, contain allergen homologues but allergies to their pollens are very scarce. Nanovesicles named pollensomes carrying allergens seemed to be a widespread mechanism of allergen transport in many clinically relevant species during in vitro germination. Prado N J Immunol. 2015 pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Oleaceae trees: facts for allergy Olive tree is an important cause of Type-I respiratory allergy in South Europe, just followed by grass pollen. Ash pollen is in the middle Europe and north of Spain after birch and grass pollen. Privet Villalba M et al 2014 The flowering period is variable: olive typically occurs between April to June and ash from late December to february Berghi N, 2014 Lilac Elongation of the pollen season may drive to perennial symptoms on the allergic patients living in the Mediterranean area and in the people coming from north and middle Europe. García-Mozo et al., 2014 Urban people sensitized to Oleaceae could present allergenic reactions during the winter due to ash pollen allergens, in the spring by the olive pollen allergens and summer as consequence of the Privet or lilac flowering. Pollutants can increase the transport of pollen into the airways or alter the respiratory epithelium of patients Schiavoni G Ann Allergy Asthma Immunol. 2017 pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Olive-allergic patients present a complex and varied IgE-binding profile West EE.UU. Mediterranean area Olive pollen is the first cause of pollinosis Japan in several Spanish and Italian provinces Australia West South America South Africa IgE binding from individual sera CBS Different allergenic patterns in individual patients allergic to olive pollen pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Allergen pattern in olive pollen extract Ole e 4 Ole e 14 Ole e 9 Ole e 11 (Polygalacturonase) (β-1,3-glucanase) Ole e 8 (PME) Ole e 12 4 EF-hand (PCBER) MM Ex 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 kDa 66 45 36 29 24 Ole e 1 20 14 Ole e 15 (Cyclophilin) Ole e 2 Ole e 3 (profilin) Ole e 10 (polcalcin) (CBM43) Ole e 7 (nsLTP) Ole e 5 Ole e 6 CRISP (superoxide dismutase) Ole e 4 results a proteolytic fragment of Ole e 9 Villalba M Methods 2013 RFAM-4 Paris 2018 Dr. Mayte Villalba Expression systems to produce recombinant allergens LTPs Ole e 7 Cys-rich Ole e 1, Fra e 1, Lig v 1, Syr v 1, Sal k 5, Ole e 6, Ole e 10 YEAST Enzymes Ole e 11, Ole e 9, Sal k 1 Pichia pastoris Profilins (Ole e 2, Che a 2, Fra e 2, Sal k 4, Sin a 4) BACTERIA EF-Hand (Ole e 8, Ole e 3, PLANTS Escherichia coli Ole e 3, Ole e 8 Che a 3, Fra e 3) Arabidopsis thaliana Others Ole e 1, NtD-Ole e 9, Ole e 12 INSECT CELLS Huecas S Eur J Biochem (1999) Spodoptera frugiperda Huecas S J Biol Chem (2001) Salamanca G FEBS J (2010) Barral P J Biotechnol (2006) Ledesma A FEBS J (2006) Mas S BBA (2017) Oeo-Santos C IAAI (2018) Ole e 10 pROF RFAM-4 Paris 2018 Dr. Mayte Villalba What characteristics make a protein an allergen? Biologic function: Proteases, glycosidases Pathogenesis-related 3D-structure: thermal and proteins: PR-2 enzymatic stability Specific location High solubilility Small size and Ole e 15 Glycosylation and high concentration polymorphism Intra/inter-specie cross-reactivity pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Families of olive allergenic proteins 14 allergen families pollen allergens characterized Ole e 1 Ole e 2 Ole e 3 Ole e 5 Ole e 6 (Regulatory protein) (profilin) (pocalcin) (superoxide dismutase) (Cys-rich) Ole e 8(Ca-binding 4-EF-Hand) Ole e 7 Ole e 9 Ole e 11 Ole e 10 (Lipid-binding protein) (b-1,3-glucanase) (pectin methylesterase) (Carbohydrate-binding) Ole e 14 Ole e 12 (polygalacturonase) Isoflavone reductase) Ole e 13 (Thaumatin) Ole e 15 (cyclophilin) pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Olive allergens have important biologic functions Remodelling pollen wall Ole e 9, Ole e 11, Ole e 14, Ole e 10, Jun a 1, Sal k 1 Redox environment Ole e 12, Ole e 5, Cyn d 4, Phl p 4Enzymes Other enzymes Ole e 15, Bet v 7 Changes in the cytoskeleton Ole e 2, Fra e 2, Sal k 4, Bet v 2 Calcium sensors Ole e 8, Ole e 3, Phl p 7, Amb a 3, Che a 3 Cellular signalling Ole e 7, Pru p 3, Bet v 1 Germination activators Ole e 1, Fra e 1, Syr v 1, Lig v 1, Sal k 5, Phl p 11, Pla l 1 Pathogenesis-related proteins Ole e 9 (PR2), Ole e 7 (PR14), Bet v 1 (PR10) Thaumatin-like Ole e 13, Cry j 3, Jun a 3 CRISP Ole e 6 Many of them bind ligands Proteins (actin) Lipids and fatty acids Carbohydrates Flavonoids and hidrophobic molecules Ions (Ca, Cu) Steroids pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Molecular features of olive allergens M 1 2 3 6 7 8 9 10 11 12 Molecular Allergen Family pI Polymorphism Glycosylation mass (kDa) Ole e 1 Ole e 1-like 5.5-6.5 18.5 Yes Yes Ole e 2 Profilin 5.1 14-16 Yes No Ole e 3 Polcalcin 4.3 9.2 Moderate No Superoxide Ole e 5 5.7 16 ND ND dismutase Ole e 6 Unknown 4.2 5.8 Moderate No Purified allergens Ole e 7 LTP >9 9.9-10.3 Yes No Ole e 8 Four EF-hand 4.5 18.8 Moderate No Ole e 9 1,3-b-glucanase 4.8-5.4 46.4 Yes Yes 3 4.5 5 6 7 8.5 10 Ole e 10 CBM-43 5.8 10.8 Yes No Pectin Ole e 11 6.3-9.3 39.6 Yes Yes methylesterase Isoflavone Ole e 12 4.8-5.7 33.9 Yes No reductase Ole e 13 Thaumatin 4.8 22.0 No No Ole e 14 Polygalacturon. 5.9- 39.6 Yes Yes 6.5 Ole e 11 Ole e 15 Cyclophilin 5.0 17.5 No No Most of allergens are acidic proteins Molecular masses between 6 to 46 kDa High polymorphism degree Some are glycoproteins pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Ole e 1, a marker of genuine sensitization to Oleaceae PM Ext kDa 66 - Most abundant protein in pollen 45 - N-glycosylated protein (heterogeneous glycan) 36 29 - Specifically expressed in pollen 24 Ole e 1 20 14 van Ree R J Biol Chem 2000 pROF RFAM-4 Paris 2018 Dr. Mayte Villalba Emergent allergenic allergens from Salsola kali and Chenopodium album Relevant allergenic source in desertic areas and salt soils Chenopodium album and Salsola kali are the main Amaranthaceae pollens in Spanish provinces Sal k 5 (Ole e 1-like) pROF Dr. Mayte Villalba Effect of sequence similarity of Ole e 1–homologues to possible IgE cross-reactivity ? Ole e 1 Zea mays Chenopodium album Che a 1 Sal k 5 Salsola kali Syringa vulgaris Fraxinus excelsior Plantago lanceolata Ligustrum vulgare Phleum pratense Lolium perenne RFAM-4 Paris 2018 pROF Dr. MayteMas VillalbaS JACI (2014) Cross-reactivity among members from Ole e 1-like Gramineae Amaranthaceae Oleaceae Plantaginaceae Betulaceae Ole e 1 Fra e 1 Syr v 1 Lig v 1 Lol p 11 Che a 1 Sal k 5 Pla l 1 BB18 Ole e 1 87 90 88 27 30 32 33 29 Fra e 1 93 84 91 30 32 34 34 30 Cross-reactivity is Syr v 1 94 91 90 28 31 33 33 28 expected among % Identity % Lig v 1 93 95 95 28 31 33 33 29 members of the same Lol p 1 44 44 43 43 42 42 25 33 phylogenetic family Che a 1 49 48 50 51 54 68 24 44 Sal k 5 50 51 51 50 57 84 26 42 Pla l 1 49 50 50 52 39 36 38 29 BB18 47 47 46 48 46 59 50 40 % Similairty Identities between Oleaceae members: 85-91% Identities between Chenopodiaceae members: 68% Identities between Chenopodiaceae and Oleaceae: 30-32% Identities between Chenopodiaceae and others: 38% RFAM-4 Paris 2018 pROF Dr.