Map-based Cloning And Functional Analysis Of The Flowering Gene FTL1 In Tomato

Tomato?Solanum lycopersicum?is one of the major vegetable crops and occupies an important position in the agricultural economy and food production.Its wild ancestor,Solanum pimpinellifolium,originates from the Andes in South America and is a short-day plant.Day-neutral tomato cultivars have spread globally from the center of origin during tomato domestication,which has reduced the strength of both their long-day and short-day flowering responses.Previous studies have showed that 52-bp sequence deletion in SELF PRUNING 5G?SP5G?3'UTR in cultivars contributes to tomato day-neutral flowering by reducing the long-day?LD?response.Why floral induction of tomato cultivars is insensitive to inductive short day?SD?is still unknown.Here,a recombinant inbred lines?RILs?population that was generated by crossing S.lycopersicum cv.Moneymaker?MM?,which is a day-neutral tomato cultivar,and S.pimpinellifolium PI365967?PP?,which is sensitive to short-day,were used.Association analysis with the whole-genome sequencing data of the RILs revealed a significant signal,named qFON10.qFON10 was ultimately mapped to a 4-kb DNA region on chromosome 11 using near-isogenic lines?NILs?and BC₃F₂ individuals.Two annotated genes,Solyc11g008650 and Solyc11g008660,homologs of the Arabidopsis FT gene,were located in this region.RT-PCR amplification and sequencing confirmed that the predicted Solyc11g008650 and Solyc11g008660responded to a single gene,named Flowering Locus T-Like 1?FTL1?.The 2-bp deletion in Solyc11g008650 causes early termination of FTL1,with a fragmentary PEBP domain in MM.We further knocked out endogenous FTL1 using CRISPR/Cas9 technology,and the ftl1cr mutants were obtained.All these ftl1cr mutants displayed later flowering than wild type under short day conditions,but the flowering time of the ftl1cr mutants and wild type displayed no obvious differences under natural long day conditions,indicating that FTL1 only regulate short-day flowering in tomato.In line with this,FTL1 was induced specifically by short day conditions rather than long day conditions.Yeast two-hybrid and Bimolecular Fluorescent Complimentary showed that FTL1 interacts with 14-3-3/2 to form FAC,with SP-interacting G-BOX?SPGB?regulating tomato flowering.Genetics research indicating that SFT act downstream of FTL1 to regulate flowering under short day conditions.Different allelic combinations of SP5G and FTL1 occurred in the same region,suggesting that the de novo mutations of SP5G and FTL1occurred during initial domestication.And simultaneous mutations in SP5G and FTL1 in short-day wild tomato accession could result in day-neutral flowering.Our findings revealed the function of the FT homologous protein FTL1 in the regulation of tomato flowering,analyzes the molecular mechanism of tomato response to short-day flowering,and provided new insight into the roles of FTs in the modulation of photoperiodic flowering in plants.