Effects of expression of lcb1/lcb2 and lac1/lag1 genes on the biosynthesis of ceramides

Abstract

Ceramides are important signal messenger molecules due to their role in regulating diverse responses such as cell cycle arrest, apoptosis, and senescence. Yeasts are more suitable for ceramides production than any other microorganisms since they grow fast and are non-pathogenic. However, regulations of the expression of genes involved in sphingolipid synthesis pathway are required to enhance ceramides production. In this study, we investigated the effects of co-expression of two pairs of genes that encode for serine palmitoyltransferase and ceramide synthase, respectively. Effect of other genes of the enzymes associated with the sphingolipid synthesis pathway, 3-ketosphinganine reductase and sphinganine C-4 hydroxylase were also studied and compared. The genes were cloned in to pESC-URA vector. Saccharomyces cerevisiae was cultivated aerobically in YPDG medium at 30°C. Ceramides were seperated from cell extracts by solvent extraction and quantified by HPLC with ELSD. The highest ceramides production (10.52 mg ceramides/g cell) was obtained when 3-ketosphinganine reductase, which is encoded by tsc10 gene, was overexpressed. Also, S. cerevisiae SCEL2,1 overexpressing serine palmitoyltransferase encoded by lcb2 and lcb1 genes, and S. cerevisiae SCEG1C1 overexpressing ceramide synthase encode by lag1 and lac1 genes, showed a high level of ceramides production (10.08 mg ceramides/g cell and 9.88 mg ceramides/g cell, respectively).