The genes BnSCT1 and BnSCT2 from Brassica napus encoding the final enzyme of sinapine biosynthesis: molecular characterization and suppression

Abstract

This study describes the molecular characterization of the genes BnSCT1 and BnSCT2 from oilseed rape (Brassica napus) encoding the enzyme 1-O-sinapoyl-beta-glucose:choline sinapoyltransferase (SCT; EC 2.3.1.91). SCT catalyzes the 1-O-beta-acetal ester-dependent biosynthesis of sinapoylcholine (sinapine), the most abundant phenolic compound in seeds of B. napus. GUS fusion experiments indicated that seed specificity of BnSCT1 expression is caused by an inducible promoter confining transcription to embryo tissues and the aleurone layer. A dsRNAi construct designed to silence seed-specifically the BnSCT1 gene was effective in reducing the sinapine content of Arabidopsis seeds thus defining SCT genes as targets for molecular breeding of low sinapine cultivars of B. napus. Sequence analyses revealed that in the allotetraploid genome of B. napus the gene BnSCT1 represents the C genome homologue from the B. oleracea progenitor whereas BnSCT2 was derived from the Brassica A genome of B. rapa. The BnSCT1 and BnSCT2 loci showed colinearity with the homologous Arabidopsis SNG2 gene locus although the genomic microstructure revealed the deletion of a cluster of three genes and several coding regions in the B. napus genome.

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

“…In some experiments, a band at 52 kD was also detected at low levels in Fraser et al (2005). Blue circles indicate functionally characterized SCPL acyltransferase enzymes that have previously been reported: SCT, sinapoylglucose:choline sinapoyltransferases from Arabidopsis (Shirley et al, 2001) and B. napus (Weier et al, 2008); SMT, Arabidopsis sinapoylglucose:malate sinapoyltransferase (Lehfeldt et al, 2000); SAT, Arabidopsis sinapoylglucose:anthocyanin sinapoyltransferase (Fraser et al, 2007); SST, Arabidopsis sinapoylglucose:sinapoylglucose sinapoyltransferase (Fraser et al, 2007); GAC, glucose acyltransferase from wild tomato (Li and Steffens, 2000). Oat SCPL1 (SAD7) is indicated by the red circle.…”

“…Since some dicot SCPL proteins are known to have acyltransferase functions (Lehfeldt et al, 2000;Li and Steffens, 2000;Shirley et al, 2001;Fraser et al, 2007;Weier et al, 2008), and Sad7 is known to be genetically linked to Sad1 and Sad2, SCPL1 became a candidate avenacin acyltransferase gene. We sequenced the SCPL1 gene in mutants #376 and #616 and identified a single nucleotide polymorphism (SNP) in each.…”

“…Most of these are either from Arabidopsis thaliana or Brassica napus and fall within a Brassicaceae-specific subgroup of the serine carboxypeptidase/ SCPL protein family within Clade 1A (Fraser et al, 2005). These enzymes include the sinapoylglucose:malate sinapoyltransferase from Arabidopsis (SNG1), which is required for the synthesis of the UV protectant sinapoylmalate (Lehfeldt et al, 2000), and sinapoylglucose:choline sinapoyltransferases (SCTs) from Arabidopsis (SNG2) and B. napus (Bn SCT1 and Bn SCT2), which are required for the formation of sinapoylcholine in seeds (Shirley et al, 2001;Weier et al, 2008). The wild tomato (Solanum pennellii) SCPL acyltransferase GAC, which catalyzes the formation of glucose polyesters associated with insect resistance in wild tomato (Li and Steffens 2000), does not lie within the Brassicaceae-specific subgroup but does group within Clade 1A (Fraser et al, 2005).…”

“…Members of the BAHD family are involved in the synthesis of important natural products, including the anticancer drugs vinblastine (St Pierre et al, 1998) and taxol (Kingston 1994;Walker et al, 2002). More recently, serine carboxypeptidase-like (SCPL) proteins have emerged as a new group of acyltransferase enzymes that are able to modify plant natural products (Lehfeldt et al, 2000;Li and Steffens, 2000;Shirley et al, 2001;Fraser et al, 2007;Weier et al, 2008). These two different classes of acyltransferases operate by distinct mechanisms.…”

“…The SCPL acyltransferases that have been cloned and characterized to date are all of dicot origin (Lehfeldt et al, 2000;Li and Steffens, 2000;Shirley et al, 2001;Fraser et al, 2007;Weier et al, 2008). Oats (monocots belonging to the genus Avena) produce acylated defense compounds known as avenacins.…”