PROJECT SUMMARY/ABSTRACT The overarching goal of this AREA project is to dissect the molecular mechanisms and evolution of glyceollin variation toward the goal of successful and efficient metabolite engineering to produce optimal levels of glyceollins for human health and cancer treatment. Glyceollins are induced phytoalexins that are naturally synthesized in legumes (such as soybeans and wild soybeans) in response to infections or stress. Glyceollins are members of the isoflavonoid family, and they exhibit antioxidant and anti-inflammatory activities as well as an anti-estrogenic activity, which has been shown to suppress human breast and ovarian carcinoma tumorigenesis. Studies have focused on the medical properties of glyceollins, but little is known about glyceollin variation and genetic determination in natural plant populations. Knowledge about the evolutionary factors and processes that play important roles in maintaining these complex trait variations is also limited. This lack of fundamental knowledge about these important bioactive compounds limits the success of plant metabolite engineering to produce optimal levels of glyceollins needed for cancer treatment and other chronic diseases. This project is the first to examine the molecular mechanisms and evolution underlying the complex variation of glyceollins in natural populations of soybeans integrating multiple disciplines, such as plant genomics, transcriptomes, biochemistry, and evolutionary genetics. The research goal will be accomplished by completion of the following 3 specific aims: 1) To assess the genetic variation of stress-induced glyceollins among diverse wild soybean populations; 2) To dissect the genetic basis of induced glyceollin variation in natural populations using RNA-seq based transcriptome comparisons and genome-wide association studies; and 3) To validate the function of the most promising candidate genes using a soybean root transformation system. The research outcomes will improve the scientific knowledge of the molecular basis of glyceollin natural variation and the evolutionary processes that maintain these variations. The comprehensive transcriptome and genomic data will provide a community resource for generating and testing new hypotheses and for deployment of similar strategies to the study of other human health-related complex traits. This AREA award will enhance the research environment in the Department of Biological Sciences at the University of North Carolina at Charlotte, a minority-serving institution, by providing hands-on experiences for graduate and undergraduate students in the PI's lab. Additionally, it will be integrated with UNC Charlotte's effort in the Plant Pathways Elucidation Project (P2EP), an education and research program that teams university scientists, industry leaders and college students from across North Carolina to investigate how plants can benefit human health.