Yezhang Ding, Alisa Huffaker, Tobias G. Köllner, Philipp Weckwerth, Christelle A.M. Robert, Joseph L. Spencer, Alexander E. Lipka, Eric A. Schmelz

To ensure food security, maize (Zea mays) is a model crop for understanding useful traits underlying stress resistance. In contrast to foliar biochemicals, root defenses limiting the spread of disease remain poorly described. To better understand below-ground defenses in the field, we performed root metabolomic profiling and uncovered unexpectedly high levels of the sesquiterpene volatile β-selinene and the corresponding non-volatile antibiotic derivative, β-costic acid. The application of metabolite-based quantitative trait loci (mQTL) mapping using bi-parental populations, genome wide association studies, and near-isogenic lines (NILs) enabled the identification of terpene synthase 21 (ZmTps21) on chromosome 9 as a β-costic acid pathway candidate gene. Numerous closely examined β-costic acid deficient inbred lines were found to harbor Zmtps21 pseudo genes lacking conserved motifs required for farnesyl diphosphate (FPP) cyclase activity. For biochemical validation, a full length ZmTps21 was cloned, heterologously expressed in E. coli and demonstrated to cyclize FPP yielding β-selinene as the dominant product. Consistent with microbial defense pathways, ZmTps21 transcripts strongly accumulate following fungal elicitation. Challenged field roots containing functional ZmTps21 alleles displayed β-costic acid levels over 100 μg g-1 FW, greatly exceeding in vitro concentrations required to inhibit the growth of five different fungal pathogens and rootworm larvae (Diabrotica balteata). In vivo disease resistance assays, using ZmTps21 and Zmtps21 NILs, further support the endogenous antifungal role of selinene-derived metabolites. Involved in the biosynthesis of non-volatile antibiotics, ZmTps21 exists as a useful gene for germplasm improvement programs targeting optimized biotic stress resistance.