A ShbHLH34‐ShPUR Module Activates ShTPS1‐mediated β‐caryophyllene Biosynthesis for Insect Resistance in Sugarcane

Terpenoids represent one of the largest and most structurally diverse classes of plant secondary metabolites, playing essential roles in defense against herbivorous insects. Despite their importance, the mechanisms regulating terpenoid biosynthesis remain largely unexplored. Here, in sugarcane, we identified and characterized a terpene synthase gene from Saccharum spp. Hybrids, ShTPS1, which is strongly induced by feeding from fall armyworm Spodoptera frugiperda and sugarcane stem borer Chilo infuscatellus. Enzymatic assays in vitro revealed that ShTPS1 catalyzes the biosynthesis of volatile terpenoid β-caryophyllene using substrate farnesyl diphosphate (FPP) and β-caryophyllene showed a strong repellent effect on S. frugiperda, Chilo infuscatellus larvae and Ceratovacuna lanigera. ShTPS1 overexpression in vivo in tobacco and sugarcane both significantly enhanced β-caryophyllene emission and dramatically reduced larval feeding preference. We further revealed that the basic helix-loop-helix transcription factor ShbHLH34 activates ShTPS1 transcription by binding to a G-box motif in its promoter. Notably, this ShbHLH34-mediated ShTPS1 transcription was significantly enhanced by a transcriptional cofactor purine-rich alpha ShPUR. Collectively, our findings establish the ShbHLH34/ShPUR-ShTPS1 regulatory module as a key mechanism underlying herbivore-induced terpenoid biosynthesis in sugarcane and provide promising molecular targets for breeding insect-resistant varieties through genetic improvement.