br Introduction br Sphingolipid metabolism Sphingolipids are
Sphingolipid metabolism Sphingolipids, are fundamental constituents of all eukaryotic membranes and their metabolism is carried out by a broad array of anabolic and Digoxin synthesis with ceramide as their hub (for a review, (Hannun and Obeid, 2018)). Ceramide can be formed by multiple pathways: the de novo pathway where ceramide is synthesized from non-sphingolipid precursors, the multi-step hydrolysis of complex sphingolipids and the salvage pathway where sphingosine can be re-acetylated to form ceramide. The de novo biosynthesis of sphingolipids begins with the condensation of serine and palmitoyl-CoA, a reaction catalyzed by the ER-located enzyme serine palmitoyltransferase (SPT) (reactions are depicted in Fig. 1). Its product, 3-ketodihydrosphingosine, is converted to dihydrosphingosine by the enzyme 3-ketodihydrosphingosine reductase (KDHR). Ceramide synthases (CerS1-6) N-acetylate the dihydrosphingosine backbone to form dihydroceramide, which can be then transformed to ceramide by the enzyme dihydroceramide desaturase (DES). Interestingly, ceramide can be further modified at the 1-hydroxyl position to form more complex sphingolipids (Fig. 1). For instance, ceramide kinase (CERK) catalyzes its phosphorylation to generate ceramide-1-phosphate. Sphingomyelin is the product of the sphingomyelin synthases activity (SMS1-2), which transfer a phosphocholine moiety from phosphatidylcholine to the 1-O-position. Glucosylceramides are produced by the addition of UDP-galactose (galactosyltransferase, CGT) or UDP-glucose (glucosylceramide, GCS) at the ceramide backbone. More recently, we discovered the formation of O-acylceramides by the addition of a fatty acyl (FA) chain at the 1 or 3-O-hydroxyl and catalyzed by the ceramide O-acyltransferase activity of diacylglycerol acyltransferase, DGAT 1 or 2 enzymes (see in detail below) (Senkal et al., 2017). Interestingly, CerS1-6, fatty acyl-CoA synthase (ACSL) and DGAT2 were found to form a multi-enzyme complex on the ER- LD interface (Senkal et al., 2017) (Fig. 2).
Diacylglycerol acyltransferase (DGAT) enzymes Triacylglycerides (TAG) are the principal source of energy storage in eukaryotic cells. TGs are neutral lipids formed by a glycerol backbone and three molecules of fatty acids (FA) covalently linked by ester bonds. FAs need to be activated to acyl-CoA FAs by the ACSL enzyme. Because of their hydrophobic nature, TGs are stored in plasma lipoproteins or in cytosolic LDs. TGs can be synthesized within the cell by two pathways: the glycerol phosphate or Kennedy pathway and the monoacylglycerol pathway. Both pathways converge into the last reaction of TAG synthesis, where diacylglycerol (DG) is converted to TAG by the action of the acyl-CoA:diacylglycerol acyltransferase (DGAT, EC 126.96.36.199) enzyme (Yen et al., 2008). Several lines of evidence suggested that DGAT2 is involved in the bulk of TAG synthesis. For example, overexpression of DGAT2 in rat hepatoma cells showed significantly more TAG mass accumulation than DGAT1 overexpressing cells (Stone et al., 2004). Conversely, deletion of the yeast ortholog of DGAT2 (Dga1) resulted in higher reduction of TAG than the ortholog of DGAT1 (Are1 or Are2) (Sorger and Daum, 2002) (Oelkers et al., 2002). Likewise, DGAT2-deficient mice (Dgat2(−/−)) had deficient lipid body (lipopenia) and died soon after birth due to impaired permeability barrier function in the skin (Stone et al., 2004), whereas DGAT1-deficient mice (Dgat1(−/−)) showed less TAG in the liver and mammary gland (Smith et al., 2000). Mutations in DGAT1 associated with decreased levels of DGAT1 protein were identified in patients with congenital diarrheal disorders. Patient-derived organoids showed altered TAG metabolism, such as reduced TAG levels and lack of LD formation when cells were loaded with oleic acid. In addition, DGAT1-deficient cells were sensitized to lipid stress upon oleate treatment which resulted in increased caspase3/7 activation as compared to control. Overexpressed DGAT1 or DGAT2 reversed the phenotype observed in the organoids derived from mutant DGAT1 patients (van Rijn et al., 2018).