title: Targeted gene disruption coupled with metabolic screen approach to uncover the LEAFY COTYLEDON1-LIKE4 (L1L4) function in tomato fruit metabolism.
abstract: Functional analysis of tomato L1L4 master transcription factor resulted in important metabolic changes affecting tomato fruit quality. Tomato fruits from mutant lines bearing targeted disruption of the heterotrimeric nuclear transcription factor Y (NF-Y) transcription factor (TF) gene LEAFY-COTYLEDON1-LIKE4 (L1L4, NF-YB6), a master regulator of biosynthesis for seed storage proteins and fatty acids, were evaluated for metabolites content and morphology. Metabolic screens using LC-MS/MS-based analysis and physico-chemical methods in different L1L4 mutants of the fourth generation allowed a comparative assessment of the effects of the TF disruption. Mutagenesis resulted in fruits phenotypically similar to wild-type with subtle shape differences in the distal end protrusion and symmetry. Conversely, mutant fruits from independent lines had significant variation in moisture content, titratable acidity and overall metabolite profiles including oxalic and citric acid, fructose, β-carotene, total polyphenols and antioxidants. Lines 6, 7 and 9 were the richest in β-carotene and antioxidant activity, line 4 in ascorbic acid and lines 4 and 8 in succinic acid. The reduced content of the anti-nutrient oxalic acid in several mutant fruits suggests that L1L4 gene may regulate the accumulation of this compound during fruit development. Detailed LC-MS/MS analysis of mutant seeds showed substantial differences in bioactive compounds compared to wild-type seeds. Taken together, the results suggest that the L1L4 TF is a significant regulator of metabolites both in tomato fruit and seeds providing a molecular target for crop improvement. Elucidation of the candidate genes encoding key enzymes in the affected metabolic pathways aimed to facilitate the L1L4 gene network exploration and eventually lead to systems biology approaches in tomato fruit quality.
authors.affiliation: Meditbio, FCT, University of Algarve, Edf. 8 Campus de Gambelas, 8005-139, Faro, Portugal.
authors.forename: Custódia
authors.initials: C
authors.lastname: Gago
authors.affiliation: Institute of Applied Biosciences, Centre for Research and Technology Hellas, 6th klm Charilaou-Thermi Rd., 57001, Thessaloniki, Thermi, Greece.
authors.forename: Victoria
authors.initials: V
authors.lastname: Drosou
authors.affiliation: Department of Agriculture, Technological Educational Institution of Crete, 710 04, Heraklion, Crete, Greece.
authors.forename: Konstantinos
authors.initials: K
authors.lastname: Paschalidis
authors.affiliation: Meditbio, FCT, University of Algarve, Edf. 8 Campus de Gambelas, 8005-139, Faro, Portugal.
authors.forename: Adriana
authors.initials: A
authors.lastname: Guerreiro
authors.affiliation: Meditbio, FCT, University of Algarve, Edf. 8 Campus de Gambelas, 8005-139, Faro, Portugal.
authors.forename: Graça
authors.initials: G
authors.lastname: Miguel
authors.affiliation: Meditbio, FCT, University of Algarve, Edf. 8 Campus de Gambelas, 8005-139, Faro, Portugal.|CEOT, FCT, University of Algarve, Edf. 8 Campus de Gambelas, 8005-139, Faro, Portugal.
authors.forename: Dulce
authors.initials: D
authors.lastname: Antunes
authors.affiliation: Institute of Applied Biosciences, Centre for Research and Technology Hellas, 6th klm Charilaou-Thermi Rd., 57001, Thessaloniki, Thermi, Greece. zhilioti@certh.gr.
authors.forename: Zoe
authors.initials: Z
authors.lastname: Hilioti
mesh_terms: D005638:Fruit / Q000235:genetics / Q000378:metabolism*; D018506:Gene Expression Regulation, Plant / Q000235:genetics / Q000502:physiology; D018551:Solanum lycopersicum / Q000235:genetics / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D010940:Plant Proteins; D014157:Transcription Factors
keywords: Fruit quality; L1L4/NF-YB6; Metabolites; Tomato; Transcription factor; Zinc finger nuclease technology