## 5 Biosynthesis of Indole Alkaloids from the Stigonematales
Notably, no studies have, to date, elucidated the biosynthetic origin of the haplindolinones (Group 3), although it is assumed to occur via oxidation (to form the oxindole ring), and subsequent rearrangement (to form the spiro-fused cyclopropane ring). Prenylation of C-2 of the indole ring (of a tetracyclic haplindole precursor) would produce tetracyclic ambiguities from Group 4. And if the isoprene group reacts further, intermolecular cyclization would lead to the pentacyclic ambigunes and fischambiguines of Group 5 and 6. Oxidation, and various rearrangement of these compounds, would subsequently afford variants within the ambiguities sub-classes. Figure 14: Biosynthesis of indole alkaloid sub-classes from a tricyclic precursor. The scheme shown is specifically based on a haplindole C (_i.e._, isonitrile-bearing) precursor. The second pathway would support cyclization of the tricyclic hapalindole between C-2 of the indole ring, and C-16 of the isopropylidene group, producing the tetracyclic fischefindoles of Group 7. The condensation of the hapalindole would presumably occur via acid catalysis, and be possibly governed by an enzymatic reaction. [PERSON] _et al._[PERSON] _et al._ (2013) showed that the cyclization reaction was possible without the presence of an enzyme using hapalindole C formamide as a starting product. A strong acid was used to convert hapalindole C formamide to fischeindole C formamide and fischeindole C amine ([PERSON] _et al._, 1988). Using the same conditions, however, this same reaction was not reproducible with hapalindole E formamide that only differs from hapalindole C formamide by presence of a chlorine atom. Recent identification of a halogenase [PERSON] _et al._ (2013) capable of utilizing fischeirindole as a substrate, however, suggests chlorination could occur post-cyclization. Moreover, the identification of novel fischeirindoles (Figure 10) with 6-membered \"B-rings\" suggest an alternative, but yet investigated, biosynthetic route. Finally, a third pathway suggests the oxidation of the tricyclic hapalindole to produce an intermediate indolinone with a structure essentially similar to a tricyclic variant of anhydroahpaloxindole A. Anhydrohapaloxindole A has been shown to be produced from singlet oxygen oxidation of hapalindole A ([PERSON] _et al._, 2013; [PERSON] _et al._, 2013), and similar oxidation of a tricyclic hapalindole could presumably occur. According to the biosynthesis proposed by [PERSON] _et al._[PERSON] _et al._ (2014) this intermediate would then undergo intermolecular cyclization between C-3 of the indole ring, and C-16 of the isopropylidene group, via acid catalysis to produce wellwitindolinone A from Group 8. In order to form the remaining wellwitindolinones from Group 9, it was proposed that an isocyano epoxide intermediate would be formed, and further intermolecular rearrangements would occur. Subsequent, oxidation and methylation of the wellwitindolinones would, in particular, produce variants such as the oxidized wellwitindolinones including 3-hydroxy-N-methylwellwitindolinone C isonitrile. That said, it is worth noting that more recent assessments have proposed an alternative pathway for the wellwitindolinones, and specifically oxidative ring contraction of 12-_epi_-fischeirindole I to give wellwitindolinone A ([PERSON] _et al._, 2013), however, this has yet to be investigated. As previously mentioned, only very recently have relevant gene clusters linked to the biosynthesis of the indole alkaloids been elucidated ([PERSON] _et al._, 2014; [PERSON] _et al._, 2013; [PERSON] _et al._, 2013; [PERSON] _et al._, 2013). Work by [PERSON] _et al._[PERSON] _et al._ (2014); [PERSON] _et al._ (2013) identified gene clusters from _F. ambigua_ and _H. wellwitschi_, and established a link to the biosynthesis of ambiguine and wellwitindolinone alkaloids, respectively.