Position-specific isotope labelling gives new insights into chiral monoterpene synthesis of Norway spruce (Picea abies L.)

Chiral forms of monoterpenes and their enantiomeric composition are of ecological significance, e.g. for plant-insect interactions. However, biosynthetic pathways and drought-induced changes of enantiomeric monoterpene emissions are barely understood. We analyzed, for the first time, drought effects on the enantiomeric composition of de novo vs. storage emitted monoterpenes from Norway spruce saplings by position-specific ¹³C-pyruvate (¹³C2- and ¹³C1-labelled) feeding and ¹³CO₂ fumigation. Drought reduced total monoterpene emissions already during its early stages, strongly linked to net photosynthesis, and lead to a decline in de novo synthesis of monoterpenes. However, it unevenly affected chiral monoterpenes, leading to compositional changes of emissions with increasing drought. At the onset of drought, the (-)-enantiomers of limonene, β-phellandrene, α- and β-pinene were emitted at higher rates than the (+)-enantiomers. Our results suggest that (-)-enantiomers were emitted mainly from storage pools while emissions of (+)-enantiomers rather depended on de novo biosynthesis. Even though biosynthesis of different monoterpenes derives from the same precursor pool, isotopic label incorporation revealed three groups among monoterpenes: storage derived, dominantly labelled via ¹³C2-pyruvate, and dominantly labelled via ¹³CO₂-fumigation. Our results contribute to a growing amount of evidence of high flexibility in metabolic pathways of monoterpene biosynthesis in plant cells.