Enzymes of B-Ring-Deoxy Flavonoid Biosynthesis in Elicited Cell Cultures of "Old Man" Cactus (Cephalocereus senilis)

Abstract

Elicited cell cultures of the cactus Cephalocereus senilis produce a group of flavonoids with unsubstituted B-rings, including an aurone which represents a new class of phytoalexin. Preliminary enzymological studies indicated that the chalcone synthase (CHS) and chalcone isomerase (CHI) from cultures of C. senilis were active with cinnamoyl-CoA and 2',4',6'-trihydroxychalcone, respectively, probable intermediates for synthesis of flavonoids with unsubstituted B-rings. We now demonstrate that the cultures contain two isoforms of CHI, both of which are induced by elicitor treatment and are active with both 2',4,4',6'-tetrahydroxy- and 2',4',6'-trihydroxychalcone. (Hydroxy)-cinnamate:CoA ligase in the cactus cultures was active with cinnamic, 4-coumaric, caffeic, ferulic, and 4-methoxycinnamic acids, but not sinapic acid. A single form of CoA ligase, as resolved by chromatofocusing analysis, was active against both cinnamate and 4-coumarate. Cinnamic acid 4-hydroxylase (CA4H) activity was induced by elicitor treatment. Thus, elicited cultures contain the necessary enzymatic activities for synthesis of B-ring-hydroxy and -deoxy flavonoids. Synthesis of only the deoxy class in response to elicitation may result from some form of metabolic compartmentation through which the CA4H reaction is bypassed, leading to formation of cinnamoyl CoA which may then be incorporated into B-ring deoxy flavonoids via nondiscriminating CHS and CHI activities.

A specialized flavone biosynthetic pathway has evolved in the medicinal plant, Scutellaria baicalensis

“…A broad-specificity 4CL activity supporting deoxyflavonoid biosynthesis was described in C . senilis ( 30 ), suggesting that a pathway different from the one we have characterized in S . baicalensis operates in this widely diverged species.…”

“…benthamiana showed that SbCLL-7 can compete with the endogenous C4H activity for cinnamic acid, an activity hypothesized, but not found, in old man’s cactus ( Cephalcereus senilis ) by Liu et al . ( 30 ). Indeed, transcript levels of C4H were particularly high in the roots and stems of Scutellaria (fig.…”

“…In Arabidopsis , there are four 4-CoA ligase isoforms that exhibit distinct substrate specificities and may participate in different flavonoid metabolic pathways ( 27 , 28 ). It has been reported that 4CL-like (CLL) proteins may activate cinnamic, benzoic, or fatty acid derivatives, although specificity for cinnamic acid has not yet been demonstrated for any of these 4CL isoforms, which tend to show similar affinity for cinnamic acid and 4-coumaric acid as substrates in vitro ( 20 – 22 , 29 , 30 ).…”

Colocalization of l-Phenylalanine Ammonia-Lyase and Cinnamate 4-Hydroxylase for Metabolic Channeling in Phenylpropanoid Biosynthesis

“…For example, some pathways of phenylpropanoid biosynthesis can bypass the C4H reaction. These include 2-hydroxylation of cinnamic acid by chloroplastic cinnamate 2-hydroxylase (Gestetner and Conn, 1974), glucosylation of cinnamic acid by UDP glucose:trans-cinnamic acid glucosyltransferase (Shimizu and Kojima, 1984;Edwards et al, 1990), and the biosynthesis of B-ring deoxy-flavonoids such as baicalein via a coumarate CoA ligase isoform that is active with cinnamic acid (Liu et al, 1995). Lignin, flavonoid, and chlorogenic acid biosynthesis in tobacco might operate through the channeled PAL isoform.…”

Cloning and expression analysis of peanut (Arachis hypogaea L.) CHI gene

“…Type I CHI could catalyze 6-hydrox-chalcone into (2S)-flavonoid or (2S)-5-desoxidation flavonoid. Type I CHI was found in most of plant species such as barley, rape, Arabidopsis, and rice (Liu et al 1995;Druka et al 2003). Type II CHI, which was mainly found in leguminous plants could catalyze both 6-hydrox-chalcone and 6-deoxidation-chalcone into (2S)-flavonoid or (2S)-5-desoxidation flavonoid.…”