Integrated multi-omics profiling reveals a landscape of dramatic metabolic defect in Artemisia annua

samples from tdd1 . It suggested a possible involvement of the correlated genes in the synthesis of the flavonoids that were absent in tdd1 . In summary, this study displays a systematical landscape of the transcriptional and metabolic changes between tdd1 and WT, arising from the GSTs defect


Dear Editor,
Trichomes are the specialized structures found on the surface of plants, categorized into glandular secretory trichomes (GSTs) and non-glandular trichomes based on their secondary metabolism capability [1].Artemisia annua possesses both of the two types of trichomes, i.e., non-glandular T-shape trichomes (TSTs) and peltate GSTs, the latter being the primary site for the synthesis and accumulation of the specific antimalarial component, artemisinin [2].Significant research efforts have been dedicated to elucidating the molecular mechanisms governing GST initiation and the metabolic pathways involved in artemisinin in A. annua [3,4].However, the comprehensive metabolism landscape of GSTs remains incompletely understood [5].
Here, we reported an A. annua mutant, which was accidentally discovered, exhibiting developmental defects in GSTs, named TRICHOME DEVELOPMENTAL DEFECTS 1 (tdd1) (Fig. 1a and b).Previous studies suggest that the GST cells are expected to possess denser cytoplasm indicative of secretory activity [6].However, the cells of defective GSTs in tdd1 were occupied by large vacuoles (Fig. 1c), revealing a compromised capacity for the secretion of secondary metabolites.
Because artemisinin was primarily accumulated in GSTs, we determined the contents of artemisinic acid, dihydroartemisinic acid, and artemisinin, which are the key products in the artemisinin biosynthesis pathway, in young and mature leaves of tdd1 and WT.Artemisinin, artemisinic acid and dihydroartemisinic acid were virtually undetectable in neither young nor mature leaves of tdd1 (Fig. 1d).This result demonstrated that the mutation of GSTs can lead to the obstruction of the artemisinin metabolic pathway.
To further uncover the metabolic difference between tdd1 and WT, young and mature leaves were collected for LC-MS based nontargeted metabolites analysis and GC-MS based volatile organic compounds (VOCs) analysis by MetWare (Wuhan, China) as described previously [7].A total of 836 distinct nontargeted metabolites, classified into 10 classes, were detected (Fig. 1e).Among these, 52 metabolites were undetectable in both YL and ML of tdd1, primarily comprising to f lavonoids (15) and terpenoids (14) (Fig. 1f).In the past decade, most studies have focused on the accumulation of artemisinin in GSTs, neglecting the potential effects of f lavonoids [8].Therefore, the nontargeted metabolome data can expand our understanding of the potential GST-specific f lavonoids in A. annua.Accordingly, 131 VOCs including 11 classes were identified (Fig. 1e).There were 38 VOCs (mainly terpenes) undetected, in both YL and ML of tdd1 (Fig. 1f).Apparently, according to our data, GST could be a specific site for the biosynthesis of many secondary metabolites, especially terpenes and f lavonoids.
of GSTs in A. annua.The synthesis of volatile terpenes shows a strong correlation with the MEP pathway [10].Therefore, the loss of the volatile terpenes in tdd1 may lead to a metabolic inhibition in the MEP pathway, which corresponds to the down-regulation of the genes of the MEP pathway.Otherwise, the complex and variable metabolic crosstalk between the MEP and MVA pathways might cause the upregulation of the gene of MVA pathways.In brief, tdd1 is an excellent mutant material to uncover the related mechanism.
To elucidate the variations in terpenes/terpenoids accumulation, we investigated the expression profile of the genes involved in the other terpenes/terpenoids biosynthesis pathways (Fig. 1h).Genes, including CPS (AA493140), GAS (AA450520, AA493970), BAS (AA329590, AA329600), LAS (AA257900, AA408270), CIN (AA458740), and CPS (AA068310, AA251590) showed dramatically low expression levels in tdd1.Integrated analysis of metabolomic and transcriptomic results suggested a substantial impediment in terpenes/terpenoids metabolism was largely hampered in tdd1.The genes with low expression levels, which exhibited the same pattern as the GST-specific genes in the artemisinin biosynthesis pathway, may play a crucial role in the synthesis and accumulation of GST-specific terpenes/terpenoids.
Since f lavonoids constituted the majority of undetectable metabolites in tdd1, we further investigated the expression levels of the enzymes involved in f lavonoid biosynthesis pathways.As a result, 33 DEGs were identified and changed in varying degrees between tdd1 and WT (Fig. 1i).Notably, genes such as 4CL (AA605090), CHS (AA325220), CHI (AA107840) and FLS (AA061500) exhibited extremely low expression levels in all samples from tdd1.It suggested a possible involvement of the correlated genes in the synthesis of the f lavonoids that were absent in tdd1.
In summary, this study displays a systematical landscape of the transcriptional and metabolic changes between tdd1 and WT, arising from the GSTs defect, and identifies specific genes that conduce to the disparate metabolites' accumulation, thereby laying the foundation for future investigations on the contributions of these genes to the GSTs-specific terpenes/terpenoids and f lavonoids biosynthesis.