Biosynthesis of ecdysteroids in in vitro prothalli cultures of Polypodium vulgare

Reixach, Natàlia; Irurre-Santilari, Josep; Casas, Josefina; Melé, Enric; Messeguer, Joaquima; Camps, Francisco

doi:10.1016/0031-9422(96)00336-6

Cited by 17 publications

(15 citation statements)

References 14 publications

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“…In our previous work with P. vulgare prothalli, we showed that this material produces significant amounts of labelled 25‐deoxy‐20,24‐dihydroxyecdysone and 25‐deoxy‐20,26‐dihydroxyecdysone after 1 week of incubation with labelled mevalonate [21]. We suggested that 25d20E could be a putative precursor of these ecdysteroids, although it was detected in minor amounts in this tissue.…”

Section: Discussionmentioning

confidence: 71%

“…Ponasterone A (25d20E) is one of the minor components of P. vulgare [21] and could be the precursor of the 25‐deoxyecdysteroids in this plant. When this precursor was incubated with prothalli and calli, it was converted into 20E at different rates.…”

Section: Resultsmentioning

confidence: 99%

“…Previous experiments had shown that [4‐ 14 C]cholesterol is converted into ecdysteroids after a one‐week incubation with prothalli [21]. It was of interest to compare the efficiency of conversion of cholesterol and various available hydroxylated derivatives (22‐hydroxycholesterol and 25‐hydroxycholesterol).…”

Section: Resultsmentioning

confidence: 99%

“…Using radiolabelled compounds, we found that the seven ecdysteroids present in these tissues, i.e. E, 20E, 24‐hydroxyecdysone, abutasterone (20,24‐dihydroxyecdysone), polypodine B (5,20‐dihydroxyecdysone), inokosterone (25‐deoxy‐20,26‐dihydroxyecdysone) and pterosterone (25‐deoxy‐20,24‐dihydroxyecdysone), could be produced from biosynthetic precursors such as mevalonate and cholesterol, whereas all 25‐hydroxylated ecdysteroids may be produced, in part, but not exclusively, from E [21]. We recently demonstrated that ecdysteroid biosynthesis in prothalli can be greatly stimulated by immersing these tissues in hot water (45 °C) [22].…”

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confidence: 99%

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Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Reixach

Lafont

Camps

et al. 1999

European Journal of Biochemistry

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Incubation of calli and prothalli of Polypodium vulgare with different tritium-labelled ecdysteroids has led to modification of some previous assumptions about the biosynthesis of ecdysteroids in plants. Thus,-hydroxyecdysone was transformed efficiently in both tissues into 20-hydroxyecdysone (20E), but no 25-deoxyecdysteroids such as pterosterone and inokosterone were formed. Likewise, incubation of 2-deoxyecdysone (2dE) produced exclusively ecdysone (E) and 20E, indicating a high 2-hydroxylase activity in both tissues, despite calli not producing phytoecdysteroids. This 2-hydroxylation was also evident in the transformation of 2,22-dideoxyecdysone (2,22dE) into 22-deoxyecdysone (22dE). Different ecdysteroids that do not occur in P. vulgare were formed in the incubation of 3-dehydro-2,22,25-trideoxyecdysone (3D2,22,25dE) by 3a-reduction and 3b-reduction and 25-hydroxylation processes. The fact that 22,25-dideoxyecdysone and 22dE were the only 2-hydroxylated products formed in this case suggests that only compounds bearing a 3b-hydroxyl group are substrates for the 2-hydroxylase. Surprisingly, 22-hydroxylation was never observed with either 2,22dE or 3D2,22,25dE, raising the possibility that it could occur at an early step in the biosynthetic pathway. In this respect, labelled 22R-hydroxycholesterol was efficiently converted into E and 20E, whereas 22S-hydroxycholesterol was not transformed into ecdysteroids, because of its unsuitable configuration at C22. Finally, the conversion of 25-hydroxycholesterol into E and 20E was greatly enhanced after thermal treatment of prothalli which induces the release of previously stored ecdysteroids. Thus, P. vulgare prothalli and calli appear to be particularly suitable models for the study of ecdysteroid biosynthesis and its regulation in plants.Keywords: 20-hydroxyecdysone; fern; hydroxycholesterol derivatives; in vitro cultures; induction.Many plant species produce C 27 ±C 29 ecdysteroids of a wide structural diversity [1±5]. These analogues of insect molting hormone (20-hydroxyecdysone, 20E) are secondary metabolites which are expected to provide some protection to plants against non-adapted phytophagous insect species [6]. Although the presence of such compounds in plants was demonstrated more than 30 years ago, little has been learnt since about their biosynthetic pathway(s), and whether it operates in a sequence that is similar to or different from that in Arthropods is not known.In vivo and in vitro labelling experiments have established that these compounds are biosynthesized from mevalonate via C 27 , C 28 and C 29 phytosterols. Cholesterol has been proved to be the intermediate in the biosynthesis of C 27 phytoecdysteroids in Taxus baccata [7]. In biosynthetic studies with Ajuga reptans roots transformed with Agrobacterium rhizhogenes, 24-methylcholesterol and 24-ethylcholesterol have been proposed as intermediates for the biosynthesis of C 28 and C 29 ecdysteroids, respectively [8]. Adler & Grebenok [9] proposed lathosterol (5a-cholest-7-en-3b-ol) as the logical ...

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Section: Discussionmentioning

confidence: 71%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

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Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Reixach

Lafont

Camps

et al. 1999

European Journal of Biochemistry

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“…Polypodium vulgare is a fern with a worldwide distribution that usually grows in dark and humid areas, and produces PE, the highest content being in rhizomes (up to 0.4% dry weight). Different culture lines of prothalli and calli from this plant have been obtained and their PE content has been described by our group [5–10]. Among those models, a calli line with undetectable levels of PE, but that is able to transform E into 20E, was selected for this study.…”

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confidence: 99%

The first cytochrome P450 in ferns

2005

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The fern Polypodium vulgare is a phytoecdysteroid (PE)‐producing plant. Cultures of P. vulgare prothalus produce PE, whereas prothalus‐derived callus cultures do not. However, this callus line can transform topically applied ecdysone (E) to 20‐hydroxyecdysone (20E), which is the last step in the biosynthetic pathway of the main plant PE. This hydroxylation is catalysed by a cytochrome P450 enzyme. E treatment of the callus line results in an increased amount of P450, showing a linear correspondence between the amount of P450 and in vivo E 20‐hydroxylation activity, estimated by measuring the bioconversion of E to 20E. This activity can be inhibited by molecules that bind to the P450‐heme group. E shows a P450‐substrate‐binding spectrum with microsomes that overexpress the P450 protein. Finally, a P450 protein was purified from E‐treated calli, this being the first P450 to be described in the pterydophyte group.

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ChemInform Abstract: Biosynthesis of Ecdysteroids in in vitro Prothalli Cultures of Polypodium vulgare.

et al. 1997

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Biosynthesis of ecdysteroids in in vitro prothalli cultures of Polypodium vulgare

Cited by 17 publications

References 14 publications

Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

Biotransformations of putative phytoecdysteroid biosynthetic precursors in tissue cultures of Polypodium vulgare

The first cytochrome P450 in ferns

ChemInform Abstract: Biosynthesis of Ecdysteroids in in vitro Prothalli Cultures of Polypodium vulgare.

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