Integrated metabolomic and transcriptomic profiling uncovers a citrus terpene synthase with in vivo–in vitro catalytic divergence and psyllid-attracting ecological function

Abstract

Citrus plants produce a rich diversity of volatile organic compounds (VOCs) that play important roles in fruit aroma, essential oils, and defense against biological stresses in aboveground tissues, yet volatile profiles in citrus roots remain poorly understood. Here, we employed an integrated metabolomic and transcriptomic approach to investigate VOCs and associated genes in Carrizo citrange ( Citrus sinensis × Poncirus trifoliata ) roots, by comparing control roots (CK), Rhizobium rhizogenes -induced hairy roots (HR) and those further elicited by methyl jasmonate (HR+MeJA). We identified 642 VOCs in the root samples, with terpenoids representing the most dominant chemical class. Comparative analysis of differential metabolites and genes between HR, HR+MeJA, and controls revealed terpenoids and candidate genes that were responsive to R. rhizogenes induction and MeJA elicitation. Among them, a putative terpene synthase gene, Cs2g07240 , was found to be highly transcribed in several aboveground tissues and markedly upregulated upon Huanglongbing (HLB) infection. In vitro heterologous expression verified that this gene converted GPP to linalool and FPP to trans-nerolidol. However, overexpressing this gene in Carrizo citrange hairy roots promoted linalool and several non-target terpenes but not trans-nerolidol. Furthermore, stable overexpression of Cs2g07240 in sweet orange ( C. sinensis ) increased linalool accumulation in leaves, whereas trans-nerolidol remained undetectable. Leaves of the overexpression lines were also more attractive to adults of Diaphorina citri , the insect vector of HLB. Collectively, this study unveils the terpenoid volatile repertoire in citrus roots and its modulation by R. rhizogenes and MeJA, while also highlighting the catalytic divergence of this synthase between in vitro and in planta contexts, as well as the tissue-specific complexity of terpenoid biosynthesis and ecological function.