**AWARDS ISSUED PRIOR TO JANUARY 20, 2025, WERE FUNDED UNDER PREVIOUS ADMINISTRATIONS AND MAY NOT REFLECT THE PRIORITIES AND POLICIES OF THE CURRENT ADMINISTRATION.** FOLATES (VITAMIN B9) ARE A FAMILY OF PLANT-DERIVED VITAMINS ESSENTIAL FOR HUMAN GROWTH AND HEALTH. AS WE CANNOT SYNTHESIZE FOLATE OURSELVES, ALL OUR FOLATE COMES INTO OUR BODIES THROUGH OUR DIETS.FOLATE DEFICIENCY LEADS TO ANEMIA IN ADULTS, AND FOLATE DEFICIENCY DURING PREGNANCY CAN LEAD TO FETAL NEURAL TUBE DEFECTS. DIETARY FOLATES ARE FOUND IN FRUITS, BEANS, AND MOST PROMINENTLY IN GREEN LEAFY VEGETABLES. STRATEGIES TO INCREASE FOLATE LEVELS IN PLANTS WILL PROVIDE A WIDER RANGE OF SOURCES TO ADDRESS NUTRITIONAL DEFICIENCY, QUALITY FOOD ACCESS AND PRODUCT VALUE ADDITION. PRIOR EFFORTS HAVE FOCUSED ON MANIPULATING THE METABOLIC PATHWAYS INVOLVED IN FOLATE BIOSYNTHESIS. THESE APPROACHES HAVE HAD SOME SUCCESS IN INCREASING DIETARY FOLATES, BUT PLANTS INTERNAL REGULATORY SYSTEMS KEEP THEM FROM OVER-PRODUCING FOLATE. GENES HAVE BEEN IDENTIFIED IN PLANTS THAT ARE PREDICTED TO FUNCTION IN FOLATE BINDING AND TRANSPORT. CONVERTING FOLATE, WHICHIS EASILY DEGRADED, TO MORE STABLE STORAGE FORMS MAY PROVE A MORE EFFECTIVE WAY OF INCREASING FOLATE LEVELS IN CROPS THAT WE EAT. THESE GENES EXIST IN TOMATO AND THE BROAD RANGE OF GENETIC DIVERSITY IN THIS AND MANY CROPS, SUGGEST THAT NATURAL GENETIC VARIATION MAY WELL EXIST THAT COULD BE TARGETEDIN TRADITIONAL BREEDING OR THROUGH BIOTECHNOLOGY TO YIELD CROPS WITH HIGHER FOLATE LEVELS.THIS PROJECT WILL A) DEVELOP AND TEST MODIFIED METHODS FOR MORE ACCURATE FOLATE ANALYSIS IN PLANT TISSUES THAT ARE NEEDED BY PLANT BREEDERS AND THE HUMAN NUTRITION COMMUNITY, B) CONFIRM THE FUNCTION OF FOLATE BINDING PROTEINS AS A MEANS TO ELEVATEFOLATE LEVELSIN TOMATO AND AS AN EXAMPLE OF THE POSSIBILITIES FOR OTHER FRUIT CROPS, C) TEST HOW WELL ELEVATED FOLATE IS TAKEN UP BY THE HUMAN GUT, D) DETERMINE THE VALUE OF FOLATE BINDING PROTEINS AS A MEANS TO ENHANCE FOLATE NUTRITIONAL CONTENT OF FRUIT, E) CREATE FOLATE-BIOFORTIFIED TOMATOES, A CROP IMPORTANT TO SMALL AND LARGE GROWERS ALIKE ALL VIA ENHANCED FUNDAMENTAL UNDERSTANDING OF TOMATO FOLATE METABOLISM, IN PARTICULAR THE FUNCTION OF PLANT PROTEINS INVOLVED IN FOLATE BINDING, TRANSPORT, AND STORAGE.US AND INTERNATIONAL CONSUMERS ARE THE ULTIMATE BENEFICIARIES OF THIS PROJECT AND WILL ACHIEVE GREATER FOOD AND NUTRITIONAL SECURITY THROUGH ACCESS TO A MORE DIVERSE AND ACCESSIBLE NUTRITIOUS FOOD SUPPLY. BREEDERS AND SEED PRODUCERS WILL BENEFIT FROM VALUE-ADDED PRODUCT OPPORTUNITIES AS WILL THOSE ALONG THE PRODUCTION, DISTRIBUTION, PROCESSING, WHOLESALE AND RETAIL CHAIN. AS TOMATO IS A HIGH-VALUE CASH CROP, THERE IS SUBSTANTIAL POTENTIAL FOR IMPACTING SMALL AND NICHE GROWERS IN THE US AND ABROAD. THE GENES TARGETED ARE HIGHLY CONSERVED ACROSS THE PLANT KINGDOM AND WILL LIKELY HAVE TRANSLATIONAL POTENTIAL IMPACTING FRUIT PRODUCERS AND RELATED INDUSTRIES WELL BEYOND TOMATO.