Defective in Cuticular Ridges (DCR) of Arabidopsis thaliana, a Gene Associated with Surface Cutin Formation, Encodes a Soluble Diacylglycerol Acyltransferase

Abstract

A key step in the triacylglycerol (TAG) biosynthetic pathway is the final acylation of diacylglycerol (DAG) by DAG acyltransferase. In silico analysis has revealed that the DCR (defective in cuticular ridges) (At5g23940) gene has a typical HX 4 D acyltransferase motif at the N-terminal end and a lipid binding motif VX 2 GF at the middle of the sequence. To understand the biochemical function, the gene was overexpressed in Escherichia coli, and the purified recombinant protein was found to acylate DAG specifically in an acyl-CoA-dependent manner. Overexpression of At5g23940 in a Saccharomyces cerevisiae quadruple mutant deficient in DAG acyltransferases resulted in TAG accumulation. At5g23940 rescued the growth of this quadruple mutant in the oleate-containing medium, whereas empty vector control did not. Lipid particles were localized in the cytosol of At5g23940-transformed quadruple mutant cells, as observed by oil red O staining. There was an incorporation of 16-hydroxyhexadecanoic acid into TAG in At5g23940-transformed cells of quadruple mutant. Here we report a soluble acyl-CoA-dependent DAG acyltransferase from Arabidopsis thaliana. Taken together, these data suggest that a broad specific DAG acyltransferase may be involved in the cutin as well as in the TAG biosynthesis by supplying hydroxy fatty acid. Triacylglycerol (TAG),3 an acyl ester glycerol, is the most efficient storage form of energy in eukaryotic cells (1, 2). In plants, TAG is the major component of the seed oils and is mainly deposited during seed and fruit development. Plant oils have a tremendous socioeconomic value in the food industry as well as in the production of lubricants and biofuels. They are also becoming increasingly important raw materials in the petrochemical industry as a substitute for non-renewable crude oil reserves (3). Thus, numerous conventional and molecular genetic strategies have been explored in attempts to increase the TAG content and modify the fatty acid composition of the plant seed oils. Isolation of the genes involved in oil biosynthesis offer exciting opportunities for overexpressing the desired gene of interest so as to redesign the plant oil metabolism for greater yields (3).TAG is usually synthesized by the sequential incorporation of acyl groups to glycerol 3-phosphate by substrate-specific acyltransferases to give lysophosphatidic acid and phosphatidic acid, respectively (4, 5). The synthesized phosphatidic acid is dephosphorylated to diacylglycerol (DAG). The last step in this pathway is catalyzed by acyl-CoA:DAG acyltransferase (DGAT; EC 2.3.1.20) that transfers an acyl group to DAG to form the TAG. DAG lies at the branch point between phospholipid and storage lipid biosynthesis. Therefore, much research has been focused on the acyl-CoA-dependent reaction catalyzed by the DAG acyltransferase, which is an integral endoplasmic reticulum protein (6) and has also been shown to be present in the oil bodies (7) and plastids (8). Several lines of evidence suggest that the TAG biosynthesis and DGAT activity are ...

The Formation and Function of Plant Cuticles

“…DCR encodes a protein of the BAHD acyltransferase family that localizes to the cytoplasm, and it has been proposed that it may be involved in acyl transfer of cutin monomers to form precursor intermediates or oligomeric structures (Panikashvili et al, 2009). However, DCR was later biochemically characterized and shown to possess in vitro diacylglycerol acyltransferase activity, leading to the formation of triacylglycerol (Rani et al, 2010). A role for cytoplasmic triacylglycerol intermediates in cutin biosynthesis is not consistent with any known steps in this pathway, yet DCR is clearly required for cutin biosynthesis in Arabidopsis floral organs.…”

Systems-Level Analysis of Nitrogen Starvation-Induced Modifications of Carbon Metabolism in a Chlamydomonas reinhardtii Starchless Mutant

“…acyltransferase activity in Morterialla ramanniana and has since been functionally validated (Lardizabal et al, 2001;Sanjaya et al, 2013) Cre06.g310200 possesses high similarity to a type-3 DGAT. Acyltransferase activity has been demonstrated for type-3 DGATs in several organisms, although not yet in an alga (Saha et al, 2006;Rani et al, 2010;Hernández et al, 2012). MLDP1 has been suggested to play a structural role in the algal lipid body analogous to that of oleosins in land plants (Moellering and Benning, 2010).…”

Apoplastic Diffusion Barriers in Arabidopsis

“…An absence of nanoridges has also been observed in dcr, a mutant in an acyltransferase of the BAHD protein family (Panikashvili et al, 2009). Although a biochemical study characterized DCR as a diacylglycerol acyltransferase in vitro, the role of DCR in cutin precursor biosynthesis in vivo remains elusive (Rani et al, 2010).…”