## 2 The Biosynthetic Pathway of the TIAs in _Catharanthus roseus_  
In recent decades, _C. roseus_ has been extensively studied for elucidating the complex biosynthetic pathways of TIAs and now it has become an ideal medicinal plant for in-depth investigating the complex molecular mechanisms for TIA biosynthesis and transport as well as their transcriptional regulation. All the TIAs in _C. roseus_ are derived from the central precursor strictosidine, which is a condensed product of the tryptophan pathway-derived tryptamine and the seco-iridoid pathway-derived secologanin by strictosidine synthase (STR) (Figure 1). Studies on the biosynthetic pathways of tryptamine and secologain have been carried out for years. Two enzymes have been revealed for their critical roles in tryptamine biosynthesis, anthranilate synthase (AS) and tryptophan decarboxylase (TDC) [7; 13], however, specific process remains to be determined. In seco-iridoid pathway, secologanin is finally generated through eight steps after the hydrolysis of geranyl diphosphate (GPP) to geraniol synthase (GES) [14], in such pathway, all the enzymes involved have been identified and the reversible reaction from 10-oxogeranial to 7-deoxyloganetic acid has also been illuminated, while the recent study did not found the intermediate product iridotrial in this reaction [15; 16; 17]. Subsequently, the central precursor strictosidine is converted into strictosidine aglycoside by strictosidine \(\beta\)-D-glucosidase (SGD) [18], and strictosidine aglycoside can be used to synthesize various kinds of TIAs. Crystal structure studies detected two new cathenamine reductases (CRs), namely, heteroyohimbine synthase (HYS) and tetrahydrorolastoninesyntase (THAS). HYS reduces cathenamine into ajmalicine and 19-epi-ajmalicine, while THAS is predominantly responsible for the conversion of tetrahydrorolastonine from strictosidine aglycon [19]. Alstonine and serpentine are the oxidation products of tetrahydrorolastonine and ajmaline respectively. Although the specialized oxidation enzyme has not been isolated, the oxidation conversion from ajmaline to serpentine present has been observed in plant vacuoles and is mainly accumulated in roots [20]. Vindoline is an important biosynthetic precursor of vincristine and vinblastine. Since the cytochrome P450 enzyme CYP71D1V2 tabersonine3-oxygenase (T3O) and an alcohol dehydrogenase (ADHL), tabersonine3-reductase (T3R) have been identified, the biosynthesis process from tabersonine to vindoline is well understood. However, the enzymes converting the strictosidine-aglycone into tabersonine remain to be identified. On the other hand, the whole biosynthesis of another precursor catharanthine remains to be revealed, although its putative intermediate, stemmadenine, has been identified for a long time [21]. After the biosynthesis of vindoline and catharanthine, they are coupled into dimeric alkaloids, a-3\({}^{\prime}\),\({}^{\prime}\)-anhydrovinblastine (AVLB), with the peroxidase a-3\({}^{\prime}\),\({}^{\prime}\)-anhydrovinblastine synthase (PRX1) [22]. The valuable TIAs in _C. roseus_, vinblastine and vincristine, were eventually derived from AVLB through multiple steps, which remain to be undetermined. Figure 1: Schematic biosynthetic pathways for terpenoid indole alkaloids (TIAs) in _Catharanthus roseus_. The updated TIA biosynthetic pathways are presented by incorporating the most recently published results. Question marks in red indicate unknown process or enzymes for the reactions.