A Serine Carboxypeptidase-Like Acyltransferase Is Required for Synthesis of Antimicrobial Compounds and Disease Resistance in Oats  

Abstract

Serine carboxypeptidase-like (SCPL) proteins have recently emerged as a new group of plant acyltransferases. These enzymes share homology with peptidases but lack protease activity and instead are able to acylate natural products. Several SCPL acyltransferases have been characterized to date from dicots, including an enzyme required for the synthesis of glucose polyesters that may contribute to insect resistance in wild tomato (Solanum pennellii) and enzymes required for the synthesis of sinapate esters associated with UV protection in Arabidopsis thaliana. In our earlier genetic analysis, we identified the Saponin-deficient 7 (Sad7) locus as being required for the synthesis of antimicrobial triterpene glycosides (avenacins) and for broad-spectrum disease resistance in diploid oat (Avena strigosa). Here, we report on the cloning of Sad7 and show that this gene encodes a functional SCPL acyltransferase, SCPL1, that is able to catalyze the synthesis of both N-methyl anthraniloyl-and benzoyl-derivatized forms of avenacin. Sad7 forms part of an operon-like gene cluster for avenacin synthesis. Oat SCPL1 (SAD7) is the founder member of a subfamily of monocot-specific SCPL proteins that includes predicted proteins from rice (Oryza sativa) and other grasses with potential roles in secondary metabolism and plant defense.

Biochemical analysis of a multifunctional cytochrome P450 (CYP51) enzyme required for synthesis of antimicrobial triterpenes in plants

“…CPMV-based systems previously have been used very successfully for production of structural proteins such as vaccines and antibodies (29,30). We also have shown this approach to be effective for expression of three triterpene biosynthetic enzymes (an acyltransferase, a methyl transferase, and a sugar transferase) required for avenacin acylation (31,32).…”

“…CPMV expression constructs carrying the AsbAS1 or AsCyp51H10 coding sequence were transformed into A. tumefaciens strain LBA4404 and leaves of N. benthamiana plants were infiltrated as previously described (29)(30)(31)(32). Full details of cloning procedures, including primer sequences (Table S2) Triterpene Analysis and Purification of 12,13β-Epoxy-16β-Hydroxy-β-Amyrin.…”

Gene Clusters for Secondary Metabolic Pathways: An Emerging Theme in Plant Biology

“…Unlike operons, the genes within these clusters are transcribed separately. The five plant clusters are for the synthesis of cyclic hydroxamic acids in maize (Frey et al, 1997(Frey et al, , 2003(Frey et al, , 2009von Rad et al, 2001;Jonczyk et al, 2008), triterpenes in oat (Avena sativa) and Arabidopsis (the avenacin and thalianol clusters, respectively; Papadopoulou et al, 1999;Haralampidis et al, 2001;Qi et al, 2004Qi et al, , 2006Field and Osbourn, 2008;Mylona et al, 2008;Mugford et al, 2009), and diterpenes in rice (Oryza sativa; the momilactone and phytocassane clusters; Sakamoto et al, 2004;Wilderman et al, 2004;Shimura et al, 2007;Swaminathan et al, 2009). These clusters are diverse in organization and function and all appear to have evolved independently (Field and Osbourn, 2008;Frey et al, 2009;Osbourn and Field, 2009;Swaminathan et al, 2009).…”

“…Analysis of the two rice clusters was aided by the fact that expression of these gene clusters is elicitor inducible and by the availability of rice genome sequence information. The oat avenacin cluster was initially defined using forward genetics, facilitated by a simple screen for loss of root fluorescence (Papadopoulou et al, 1999;Qi et al, 2006;Mugford et al, 2009). The thalianol cluster was predicted by searching the genome sequence of Arabidopsis for candidate gene clusters containing triterpene synthase (oxidosqualene cyclase) signature genes and then validated by functional analysis (Field and Osbourn, 2008).…”

Formation of plant metabolic gene clusters within dynamic chromosomal regions

“…However, the first gene cluster for a plant secondary metabolic pathway-for the synthesis of cyclic hydroxamic acids-was discovered in maize (Zea mays) in 1997 (10), and since then a secondary metabolic gene cluster for the synthesis of the triterpene avenacin has been discovered in diploid oat (Avena strigosa) (11)(12)(13)(14), and two clusters for the synthesis of different diterpenes (momilactones and phytocassanes) have been characterized in rice (Oryza sativa) (15)(16)(17)(18). These four clusters from cereals are all required for the synthesis of preformed or stress-induced compounds implicated in plant defense (15,16,19,20).…”