A soluble diacylglycerol acyltransferase is involved in triacylglycerol biosynthesis in the oleaginous yeast Rhodotorula glutinis

Abstract

A soluble diacylglycerol acyltransferase is involved in triacylglycerol biosynthesis in the oleaginous yeast Rhodotorula glutinis The biosynthesis of triacylglycerol (TAG) occurs in the microsomal membranes of eukaryotes. Here, we report the identification and functional characterization of diacylglycerol acyltransferase (DGAT), a member of the 10 S cytosolic TAG biosynthetic complex (TBC) in Rhodotorula glutinis. Both a full-length and an N-terminally truncated cDNA clone of a single gene were isolated from R. glutinis. The DGAT activity of the protein encoded by RgDGAT was confirmed in vivo by the heterologous expression of cDNA in a Saccharomyces cerevisiae quadruple mutant (H1246) that is defective in TAG synthesis. RgDGAT overexpression in yeast was found to be capable of acylating diacylglycerol (DAG) in an acyl-CoA-dependent manner. Quadruple mutant yeast cells exhibit growth defects in the presence of oleic acid, but wild-type yeast cells do not. In an in vivo fatty acid supplementation experiment, RgDGAT expression rescued quadruple mutant growth in an oleate-containing medium. We describe a soluble acyl-CoA-dependent DAG acyltransferase from R. glutinis that belongs to the DGAT3 class of enzymes. The study highlights the importance of an alternative TAG biosynthetic pathway in oleaginous yeasts.

Diversity and evolution of plant diacylglycerol acyltransferase (DGATs) unveiled by phylogenetic, gene structure and expression analyses

“…The putative DGAT3 (Glyma17g04650) gene was also highly expressed, suggesting a possible involvement of these sequences in TAG synthesis in this species, as was demonstrated for Arachis hypogaea (Saha et al , 2006), A. thaliana (Hernández et al. , 2012) and oleaginous yeast (Rani et al , 2013). A transcriptome analysis during Arabidopsis seed development showed that the expression pattern of DGAT1 was similar to DGAT3 (Peng and Weselake 2011), but the expression of DGAT3 was higher during late seed maturation.…”

“…This pathway occurs in the cytosol and involves the acylation of monoacylglycerol to DAG and the further acylation of DAG to TAG by the action of DGAT3 . A soluble protein with DGAT activity has also been identified in oleaginous yeast (Rani et al , 2013). In addition, another soluble acyltransferase involved in TAG syntheses (LPAAT: lysophosphatidic acid acyltransferases) has been identified in Arabidopsis (Ghosh et al , 2009).…”

“…In addition to the ubiquitous occurrence of DGAT1 and DGAT2 genes in plants, other DGAT-related genes have also been identified. A soluble DGAT (DGAT3) that participates on the cytosolic pathway of TAG synthesis was first identified in peanuts ( Arachis hypogaea ) (Saha et al , 2006), and more recently in A. thaliana (Hernández et al , 2012) and yeast (Rani et al , 2013). In addition, a bifunctional DGAT/wax ester synthase (WS/DGAT), homologous to Acinetobacter calcoaceticus WS/DGAT (Kalscheuer and Steinbuchel, 2003), was characterized in A. thaliana (WSD1) (Li et al , 2008).…”

Identification and characterization of a type-2 diacylglycerol acyltransferase (DGAT2) from Rhodosporidium diobovatum

“…Subsequently, another two members of DGAT3 gene family were discovered in Arabidopsis (Rani et al 2010) and castor bean. Recently, it was suggested that a soluble acyltransferase identified in the oleaginous yeast R. glutinis belongs to DGAT3 family (Rani et al 2013). The present study reports the identification and characterization of R. diobovatum DGAT2 (RdDGAT) in a TAG-deficient yeast strain.…”

Oleaginous yeast: a value-added platform for renewable oils

“…For example, in addition to a PDAT and a type 2 DGAT, Y. lipolytica also utilizes a type 1 DGAT for TAG biosynthesis (Beopoulos et al, 2012;Zhang et al, 2012). R. glutinis possesses a soluble type 3 DGAT that is presumably part of the 10S cytosolic triacylglycerol biosynthetic complex of this species (Gangar et al, 2001;Rani et al, 2013).…”